text:
TRANSFORM YOUR BRAIN
with the ultimate meditation tool. Experience profound levels of meditation in less time with theta binaural beats.
Meditation appears to produce changes in brain activity. It also can lower your heart rate, blood pressure, breathing rate, oxygen consumption, adrenaline levels, and levels of cortisol, a hormone released in response to stress...there are many types of meditation that can result in physiological benefits, such as guided meditation.
— Research published in 2013 on Guided Meditation from the Harvard Medical School
Results suggest that the presentation of binaural auditory beats can affect psychomotor performance and mood. This technology may have applications for the control of attention and arousal and the enhancement of human performance.
Duke University published research about binaural beats that investigated whether binaural beats within the electroencephalograph frequency range can entrain EEG activity.
Results showed that binaural beats, regardless of the presented frequency, can affect divergent but not convergent thinking.
Research published in 2013 on Binaural Beats from Frontiers in Human Neuroscience, a journal of the U.S. National Institutes of Health's National Library of Medicine, concludes that binaural beats increase creativity
Research from Washington University School of Medicine in St. Louis shows that when people's brains are in a Delta brain wave state, beta amyloid production in the brain ceases and the toxic material is cleared away. One thought is to explore the possibility of using brain entrainment technology to help treat Alzheimer's, but at this point all it is being used for is to help people meditate, relax and dream instantly and effortlessly.
A 2014 article on Brainwave Entrainment published in the Huffington Post discusses the work of Deepak Chopra and Dr. Rudi Tavi, Professor of Neurology at Harvard Medical School:
Two recent studies on subliminal messages have found that subconscious visual cues can improve athletic performance and reduce negative age stereotypes of physical ability.
The positive impact of Subliminal Messaging is discussed in Psychology Today from an article published in 2015
Brain Power is a well-researched, well-referenced and practical guide to maintaining and improving your mind as you age. It provides proof that the brain can adapt, grow and learn new skills with age. Even better, the tools provided are practical, achievable, and proven.
Kelly Howell also co-authored the book Brain Power, positively reviewed by the Mayo Clinic.
Biofeedback: Three Decades of "Little" Breakthroughs, Part 1
Excerpted from the "Mind-Body Connection," The Center for Mind/Body Medicine
Some of the most important "breakthroughs" in the field of biofeedback have come not from one definitive study but from the accumulation of numerous studies over many years. Rather than focusing only on the latest published articles, we felt it important to place the field of biofeedback in its historical context.
Back in the 1960s when experimental psychologist Neal Miller first demonstrated that the autonomic nervous system could be trained to alter some bodily processes, it was thought biofeedback would change the world. Miller's discovery uprooted the prevailing paradigm, that the autonomic, or visceral, nervous system was basically "dumb" and beyond voluntary control. At the time, some scientists predicted that biofeedback eventually would allow patients to "take a fully active and direct role in literally learning not to be sick" (Dienstfrey, 1991).
Over the next three decades, some 3,000 articles and 100 books on biofeedback were published. And although the research has not uncovered the kind of "unified field" originally hoped for, biofeedback has been shown to be an effective treatment for dozens of specific ailments. These include bronchial asthma, drug and alcohol abuse, anxiety, tension and migraine headaches, cardiac arrhythmias, essential hypertension, Raynaud's disease/syndrome, fecal and urinary incontinence, irritable bowel syndrome, muscle reeducation, hyperactivity and attention deficit disorder, epilepsy, menopausal hot flashes, chronic pain syndromes, and anticipatory nausea and vomiting associated with chemotherapy. While biofeedback has been used successfully to treat some psychological and mental disorders, it seems to work best with patients in which physiological processes are relevant (Futterman & Shapiro, 1986).
The most common forms of biofeedback today make use of instruments to "feed back" information about such bodily processes as muscle tension (EMG feedback), skin temperature (thermal feedback), brain waves (EEG feedback) and respiration. By watching the monitoring device, patients can adjust their thinking and other mental processes in order to control bodily processes. In some cases, subjects learn by trial and error what kind of thinking or behavior affects those processes. In other cases, subjects are taught specific methods, such as relaxation or imagery, which it is believed will have an impact on bodily functions.
One of the most exciting areas of biofeedback research today is the use of alpha-theta brainwave training. This therapy has proven effective in the treatment of various disorders, including chemical dependence, post-traumatic stress disorder, depression, anxiety, multiple personality, panic and eating disorders. Drs. Eugene Peniston and Paul Kulkosky's (1989, 1990, 1991) pioneering work showed that training chronic alcoholics to increase the lower-frequency alpha and theta brain waves, while controlling the higher frequency beta waves, resulted in significantly less depression, less craving for alcohol and less relapse. (The alpha brain wave has been associated with a tranquil, serene state, while the theta wave corresponds to a deeper meditative state.)
The researchers' treatment protocol includes 6-8 weeks of thermal biofeedback and autogenic training, followed by 30 sessions of evoked images of personal change and alpha-theta EEG biofeedback. The sessions are performed twice a day, five days a week.
The results of the therapy have been nothing short of remarkable--80 percent short-term effectiveness, with ongoing studies tracking the long-term results. The training resulted not only in decreases in alcoholic behavior (relapse, cravings, etc.) but changes in a wide range of dysfunctional personality factors (Peniston & Kulkosky, 1990). The training even affected blood chemistry: serum Beta-endorphin levels increased in patients who completed alpha-theta brain wave training (Peniston & Kulkosky, 1989).
What accounts for these changes? The current theories behind alpha-theta brainwave training are documented in an excellent article by Jonathan D. Cowan, Ph.D. in the Megabrain Report: The Journal of Mind Technology (Vol. 2, No. 3). Cowan suggests that the power of the imagery instructions given prior to the EEG training, in which patients rehearse new intentions and positive alternatives, should not be underestimated. "These images of personal change are experienced in a relaxed state, followed closely by the effect induced by alpha-theta biofeedback, which is usually very pleasant. This forms an association between the images and pleasant effect which is repeated 30 times throughout the course of therapy.... The power of [the] ... training may be partly due to inputting images and suggestions in such a way that they bypass the conscious mind, thereby benefiting from the lack of interference from adult disbelief and disempowerment."
Other researchers have used a treatment protocol similar to Peniston's with equally good results. A couple of clinicians have added another twist — patients do the EEG brainwave training together in a group setting. The supportive element in group process has long been a key factor in alcoholism treatment (12-step and other groups). But the use of brain wave training in a group has raised interesting questions about whether people can actually influence each other's brain waves. Certainly "mob psychology" indicates that people are capable of thinking and acting quite differently while in a group than when alone. But how that relates to brain chemistry remains a mystery.
In the mind-body self-regulation groups run by the Center for Mind-Body Medicine, participants regularly use biofeedback techniques together in the group. The program's director, Mary Lee Esty, believes the group practice has a supportive, reinforcing effect that enhances individual, at-home practice. The impact of biofeedback and mind-body self-regulation techniques within a small group setting merits further study. The Center plans to begin such a research project in 1996.
Increasingly sophisticated measurement devices may expand biofeedback's possibilities in the future. For example, a number of biofeedback experts, including Esty in Washington, D.C., are pioneering a new form of EEG training that uses light stimulation to increase the range and variability of patients' dominant brain frequencies. The treatment is proving successful with trauma victims whose brains have gotten "stuck" in a pattern of predominantly slow waves. EEG slowing is associated with symptoms such as anxiety, depression, irritability, fatigue, hyperactivity, distractibility, mood swings, confusion and disorganization.
More and more, biofeedback researchers are discovering that the key to wellness is not figuring out how to achieve a perpetual "theta state." The key is brain flexibility, or as Cowan writes, teaching [a person] "to perceive and control a number of different transitions among his own unique state-contexts, which differ among themselves in the amounts of each of these rhythms that they produce. Says Esty, "The objective is balance."
Clearly, the entire field of biofeedback is as ripe for new discovery as it was 30 years ago. According to Alternative Medicine: Expanding Medical Horizons, biofeedback deserves more broad clinical trials on large patient populations to determine its effectiveness. "A major reason why many patients like biofeedback training is that...it puts them in charge, giving them a sense of mastery and self-reliance over their illness and health," states the report to the National Institutes of Health. "Such an attitude may play a crucial role in the lower health care costs seen in patients after learning biofeedback skills."


After hours emergency mobile number (+61) 407 010 121, please sms a detailed message and which property you are staying in and await a response.
NEW STRICT LOCAL LAWS ARE IN PLACE REGARDING SHORT STAY ACCOMMODATION please see below details.
Welcome to the beautiful Mornington Peninsula, we sincerely hope you enjoy your stay. This property is registered under the Mornington Peninsula Shire Short Stay Rental Accommodation Local Law. Guests of this property must ensure they read and follow the requirements of the Code of Conduct during their stay.
By accepting this property by email, sms or phone you agree and will abide by the below terms and conditions of your stay. It is the guest's responsibility to read all details before making the booking. To maintain a good standard for our guests we require certain conditions to be complied with. We appreciate most will respect our property but the occasional abuse requires that we state the following conditions.
Note: Variances can be agreed on but only by arrangement with the owner in writing, Salt Accommodation is a booking service only, all maintenance and cleaning are handled by the owner and or their cleaning contractor.
When making a booking it is pending the owner's approval, should the owner decline the booking your deposit will be refunded.
THIS PROPERTY IS STRICTLY NON SMOKING INSIDE there is a minimum $300 charge if you smoke inside this property. Please dispose of all cigarette butts and do not throw into gardens or surrounds, cleaning up of butts will be deducted from the bond.
Loud parties, groups, bux parties and events strictly prohibited. The price charged is for domestic use quiet enjoyment only and not commercial. Accordingly this rate does not allow for the extra wear associated with functions in terms of cleaning, garbage removal, wear and tear, repairs etc. Use contrary to this may result in loss of your bond and/or additional payments or claims made against you at VCAT. DISTURBANCE to THE neighbours, including EXCESSIVE NOISE IS PROHIBITED and may result in termination of rental (EVICTION) and loss of bond. You must keep all noise to a minimum as this property is located in a residential street. The Mornington Peninsula has very strict Local laws for holiday houses, they must be abided by, any fines issued to the property owner from the council will be deducted from the bond.
PETS ARE STICTLY NOT PERMITTED AT THE PROPERTY AT ANY TIME Additional charges apply if you bring a pet without permission. For hygiene and allergy reasons we cannot allow pets for the comfort of future guests.
By accepting this booking you agree and understand that you MUST LEAVE THE PROPERTY THE WAY YOU FOUND IT, this includes washing and putting away of dishes, empty dishwasher, emptying inside bins into correct council bins, putting the council bins out on bin night, and giving the property reasonable tidy up and clean before you vacate. If you leave the property in a mess or very dirty you will be charged (out of your bond) an additional $60 per hour for any additional (over and above) general cleaning or re-arrangement of furniture, search for missing remotes or keys. Any damage will be deducted from the bond. Only a standard departure clean is included in the tariff.
Key safe location & code will be emailed to you the night before arrival, you must lock keys back into key safe at checkout charges apply if you lose the key/s, lock them inside or damage the key safe.
Both the owner of the property and booking manager are not responsible for the accommodation chosen not "meeting the expectations" of the accommodation required. All reasonable attempts should be made by salt accommodation to ensure the property is advertised with a thorough description and lots of photos. There is no refund if you change your mind. If you have any concerns about the level of cleaning upon arrival or any damage or maintenance issues please report these immediately to us by photos and sms or email within 2 hours of arrival. Feel free to send photos as evidence and we will accommodate any reasonable requests immediately.
The BOND REFUND is INSTRUCTED BY THE PROPERTY OWNER (salt accommodation are not accountable for bond refund decisions or outcomes) bond claims are only processed if the owner has provided photographic evidence, sworn statement and/or receipts to prove a valid claim, this is then assessed by salt and if deemed reasonable we will process the bond claim. The property owners/and or representative prepare and clean all their own properties and witness before and after each guest, after a guests departure, the property owner is required to advise on bond refund or claim within 30 days, if a bond claim is instructed by the property owner and disputed by the guest, the guest can write to the property address/owner directly stating their reasons of dispute. Salt accommodation is a booking agent only and do not handle bond disputes between the property owner and guest once a claim has been confirmed. To avoid a bond claim it is necessary for the guest to respect the booking terms, leave the property neat and tidy as you found it and exit on time, report any accidental damage before you vacate to avoid delays in repairs and getting the property ready for the next guest, if repairs or cleaning cause delays after your stay or future booking cancellations these costs or any rent loss to the owner are also deducted from the guests bond. Min $30 bond claim fee applies to the guest if a bond claim is made.
BOND is due 7 days before check in. The bond will be refunded around 30 days from when the owner advises us the bond can be refunded after your departure, subject to the above conditions being met and all is left the way it was found, if a guest is charged out of their bond for damage or cleaning and there is a balance of the bond (if any) the refund may take longer than 30 days. It is designed to cover any additional costs incurred including but not limited to, any breakage, damage or excess cleaning requirements, extra guests beyond those declared etc. If you fail to pay your bond on time salt accommodation may resell the booking to another guest with no Refund of deposit.
Off street parking is stipulated on your final confirmation. All other vehicles must park on the street. Please do not park on grassed areas or congest or block neighbours access.
Deposit amount is non -refundable and is required to secure the booking. Payment of the rental deposit constitutes the clients acceptance of these Terms and Conditions.
Cancellation if caused by the guest is not refundable unless the house is re-let for the entire period at the same rate. If a refund is made due to the property being re-let then a $100-$300 cancellation fee may apply (POA). Bookings through 3rd party websites have their own cancellation terms, please refer to the site for more information. SCHOOLIES BOOKINGS there is no refund if you cancel at any time unless the booking can be resold in full.
The property owner will make every effort to ensure the property is available as booked. However the owners reserve the right to make alterations to their property or cancel a booking due to unforeseen circumstances. If cancellation occurs from the property owner salt accommodation will provide a full refund and will endeavour to assist and try and relocate the booking to another salt accommodation listing (pending availability and similar pricing) salt accommodation cannot guarantee another property will be available, this is all subject to availability of other properties and acceptance by other individual property owners.
Number of Guests should not exceed the number stated on the Booking Approved Confirmation or subsequently agreed in writing or email. Fees will apply for excess guests not agreed with the owners in advance. Min $50 per night per guest.
Damage, Breakages, Theft and Loss are the guest's responsibility during their stay.
Departure – the property must be left in the same state to its condition on arrival. Check in is 4pmduring peak times or 2pm unless prior approval is made for an earlier check-in. Check-out time is 10-11am depending on back to back bookings, this will be confirmed on your booking details, to allow time to prepare property for the next guests. Alternate check in or out times must be approved before commencement of booking.
If trades people or service technicians need to be called out for an appliance malfunction or breakdown and it is determined that guest operator error or failure to follow instructions is the reason for the malfunction, guests will be required to pay the bill for the service call & repairs. We strongly encourage all guests to read provided manuals and booklets in welcome book or contact SALT OR THE OWNER if you need assistance during your stay.
Guests will indemnify the owner or manager of the property against all liability, which might otherwise attach to the owner of the property as a result of injury, damage or loss (personal and proprietary suffered by any person, where such injury, damage or loss has been caused by or contributed to by any act or omission of the guest/s or visitors). These conditions are an agreement between the parties. Any infringement permits the owner of the property to refuse a key, amend the rental or immediately terminate the tenancy.
The owner of the property shall make sure that the property is in a clean and tidy condition on the day of their guests' arrival at the property. If you find this is not the case, please contact SALT immediately by sms on the number provided above. We will contact the property owner to rectify the problem as soon as possible. However, please be advised, SALT Accommodation is not responsible in any way for the property and are contracted as a booking agent by the owner to rent their property. Should the property not be fit for occupation, you must contact us within 2 hours of arrival.
Please ensure you bring any linen required as per advertisement and your final confirmation as some properties do not include linen and/or towels.
Loss – the owner takes no responsibility for the tenant's personal property. Guests should have their own travel insurance for personal belongings and medical cover.
There is a minimum $100 call out fee (deducted from the bond) if we must attend to the property for noise, nuisance or any concerns complaints about your occupancy at the property.
Thank you for booking with SALT online accommodation service, we hope you enjoy your stay!


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RxCS modules documentation
==========================

The modules in the RxCS toolbox can be diveded into two categories:

    1. **API modules**. These modules contain API functions which can be called directly by a user. |br|
       The API function name do not begin with *'_'* (an underscore character).  |br|
       Please not that these modules may also contain non-API functions, which begin with *'_'* (an underscore character). |br| |br|

    2. **Internal RxCS modules**. These modules contain only non-API functions which are internal for  RxCS. |br|
       The non-API function name always begin with *'_'* (an underscore character).

This section contains list of API functions from the API modules modules which are present in the RxCS toolbox. |br|
For description of other functions please refer directly to a module source file.

.. warning::
   Do not use non-API functions unless you realy know what you are doing!

PLEASE NOTE: To preserve code simplicity RxCS toolbox do not use objects (classes).

1. API modules - signal generators:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

6.1 rxcs.sig.sigRandMult:
-------------------------

.. automodule:: sigRandMult
   :members: main

2. API modules - signal acquisition:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

3. API modules - CS signal processing:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

4. API modules - system analysis:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

5. API modules - main frames:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

6. API modules - auxiliary modules:
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

6.1 rxcs.console:
------------------------

.. automodule:: console
   :members: pack, progress, module_progress, module_progress_done, warning, info, bullet_info, note, param, bullet_param

7. Test scripts (rxcs/test):
~~~~~~~~~~~~~~~~

7.1 sigRandMult_test:
------------------------

.. automodule:: sigRandMult_test

.. |br| raw:: html

   <br />

redpajama_set_name: RedPajamaGithub


text:
A surrogate-based approach for post-genomic partner identification
Renuka C Pillutla1,
Ku-chuan Hsiao1,
Renee Brissette1,
Paul S Eder2,
Tony Giordano2,
Paul W Fletcher1,
Michael Lennick1,
Arthur J Blume1 &
Neil I Goldstein1
Modern drug discovery is concerned with identification and validation of novel protein targets from among the 30,000 genes or more postulated to be present in the human genome. While protein-protein interactions may be central to many disease indications, it has been difficult to identify new chemical entities capable of regulating these interactions as either agonists or antagonists.
In this paper, we show that peptide complements (or surrogates) derived from highly diverse random phage display libraries can be used for the identification of the expected natural biological partners for protein and non-protein targets. Our examples include surrogates isolated against both an extracellular secreted protein (TNFβ) and intracellular disease related mRNAs. In each case, surrogates binding to these targets were obtained and found to contain partner information embedded in their amino acid sequences. Furthermore, this information was able to identify the correct biological partners from large human genome databases by rapid and integrated computer based searches.
Modified versions of these surrogates should provide agents capable of modifying the activity of these targets and enable one to study their involvement in specific biological processes as a means of target validation for downstream drug discovery.
Modern drug discovery is concerned with identification and validation of novel protein targets from among the >30,000 genes postulated to be present in the human genome [1]. In understanding the importance of any new gene and its connection to a given phenotype, there is the need to know the immediate "neighborhood" of partners for each gene product since they are most likely involved in the action of the gene product. In this regard, there are few if any new chemical entities (NCEs) capable of regulating protein:protein interactions as either agonists or antagonists. In the past, peptides have sometimes been used to obtain information about protein:protein interactions as well as regulate their activity [2, 3]. This has most often been accomplished with libraries consisting of peptides between <15 amino acids in length. Using this approach, peptides have been identified which act as agonists and antagonists though, in most cases, these peptides have not shown any sequence homology to the natural ligand [4–6]. Clearly these peptides did not use any of the natural amino acid contacts required for binding of the growth factors to their receptors. As these peptides were functional but not compositional mimics, they lacked primary sequence information useful for identifying (by motif, sequence identity or similarity) the true biological partner. These results are not surprising since the putative contact domains between receptors and hormones are expected to be conformational and short peptides were probably unable to mimic a large three-dimensional shape. In this report, we describe a novel post-genomic approach (called Phenogenix®) that involves the use of complex and random peptide libraries of large size (up to 40 amino acids) and diversity (>1011 independent clones per library) in combination with computational analysis for garnering information on the natural biological partners and pathways [7]. We also show that the specificity of these peptides can be improved by mutation at the DNA level that has implications for phenotyping and the development of Site Directed Assays. Overall, the data indicates that the surrogate peptides, derived in this manner, can contain sequence information regarding the natural contact domains for both protein:protein and protein:non-protein interactions.
Criteria for a Partner Hit
The first stage in the computational analysis of our surrogates requires the alignment of the peptides into groups based on motifs or consensus regions. In addition, we examine each peptide for significant differences in the expected frequency of amino acids and the number of times a specific peptide sequence has been repeated. After defining our query strategy (e.g., entire surrogate sequence, motifs, etc.), we simultaneously search several public databases using programs such as Blastp, MAST (Motif Alignment and Search Tool) and Patternfind (see Materials and Methods). The output from each search is further analyzed based on criteria described in Table 1. Homology between the partner and surrogate oftentimes ranges over a long stretch (15–20 amino acids) or may be found in a perfect match within a short sequence of 5–8 amino acids (unpublished data). Other positive indicators include: 1. the appearance of the partner in at least 50% of the top cohort (i.e., first 10 matches) of any one search; 2. the appearance of the same or related hits occurring in several different searches; 3. the identification of the same partner for multiple peptides from the same or related pans. Criterion 2 addresses the biological relevance of a hit (e.g., distribution, disease indication, etc.) and criterion 3 relates to the biological activity of the surrogate and its ability to cause a phenotypic change in the appropriate test system (phenotyping).
Table 1 Criteria for a Partner Hit
Panning of mRNA targets
In one series of experiments, biotinylated oligonucleotides comprising the UTRs (untranslated regions) of four mRNAs were synthesized (Table 2). The oligonucleotides were heat denatured and allowed to anneal at room temperature to allow the appropriate re-folding. All of the mRNAs were subjected to 4 rounds of panning using both 40 mer and 20 mer random libraries under similar but not equivalent conditions. Individual phage clones from rounds three and four were amplified, tested for binding to the specific and a non-specific mRNA and sequenced. Table 3 shows the overall results that were obtained from each of the pannings. On average, about 8% of the surrogates were found to be specific for each target when compared to a control RNA (RRE). For each RNA target, the predicted amino acid sequences of the peptide binders were analyzed in terms of both overall amino acid content and the occurrence of known RNA-binding motifs and consensus domains. Two motifs were observed for the APP and HCV RNAs (Figure 1 and see below). RNA binding proteins are known to have an overall abundance of certain amino acid residues [8, 9]. Table 4 shows a comparison of the specific amino acid composition of peptide binders with regard to their average frequency of occurrence seen within the original unpanned library. All of the peptide binders showed enrichment of arginine residues, as would be expected for RNA binding proteins. Also, tryptophan, serine, and glycine residues were enriched. In addition, several peptide binders showed the presence of the RGG box (Figure 2A) and one sequence was found that contained the KH motif (Figure 2B), both of which are known RNA-binding motifs [8, 9]. The isolation of surrogates containing generic RNA binding motifs is not unexpected and probably results from enhanced binding and concomitant enrichment of these peptides during the panning process. In addition, an additional consensus motif was identified among peptides isolated by panning on RRE RNA [10]. This motif [K/R] LRRR, aligns with a region on the expected natural partner, the Rev peptide (Figure 3).
Peptide surrogates with RGG Box Sequences. Random peptide libraries were panned on four different mRNA targets. Isolated phage binders from rounds three and four of each pan, were sequenced. Several peptides from each pan showed the presence of the RGG box, a well-defined RNA-binding motif [8, 9]. RGG sequences in each surrogate is in bold and underlined. Peptide surrogate with KH Domain. Panning of the 20-mer random peptide library on target M1 isolated a phage clone containing the sequence VIGxxGxxF which is similar to an RNA-binding motif, the KH motif [8, 9]. The surrogate motif corresponding to the KH domain is in bold and underlined.
RRE Binding Motif. Alignment of Rev peptide with surrogate peptides containing the (K/R)LRRRP motif. surrogates were obtained by panning a portion of the RRE mRNA using the 40 mer random peptide library. Consensus motifs are in bold and underlined.
surrogate Maturation. One clone for each of the three targets – APP, HCV, and IGF was identified for generation of secondary libraries. Residues that were selected for after four rounds of panning are indicated in bold and underlined.
Table 2 Description of targets used for the mRNA pans
Table 3 Results of panning experiments on selected mRNA targets using the primary phage displayed peptide libraries
Table 4 Comparison of Amino Acid Frequency in surrogates with expected frequencies.
One peptide was chosen from each of the APP, HCV and IGF pans (based on the highest specific binding vs. RRE RNA) as templates for the construction of secondary libraries. Each of these libraries contained >1010 independent clones and was used for panning the appropriate target RNA. Results are shown in Figure 4 and Table 5. Since library construction was based on peptides previously selected for binding and selectivity to specific RNAs, the number of target-specific clones increased dramatically following in vitro maturation. In addition, motifs were observed suggesting the critical nature of these specific residues in terms of binding to target. Preliminary studies have shown that the secondary surrogates have higher relative affinities when compared to the original clones (unpublished data).
HCV-eIF3 Binding Motif. Alignment of eIF3 with surrogate peptides containing the TxRLL motif. surrogate peptides were obtained by panning a portion of the 5'UTR of HCV mRNA using both the 20 mer and 40 mer random libraries. Peptides HCV-3-F5 and HCV-3-H8 were obtained from the 40-mer library from the same pan. Peptide HCV-NG-D9 was obtained from the 40 mer library using modified experimental conditions. Peptide HCV-3-C3 was obtained from the 20 mer library. Consensus sequences are in bold and underlined. Sequences outside the motif that are conserved between the surrogates and eIF3 are in Italics and underlined.
Table 5 Results of panning experiments on RNA targets using secondary libraries
In the case of the HCV RNA, we panned a sequence (AA UUG CCA GGA CGA CCG GGU CCU UUC UUG GAU CAA CCC GCU CAA UGC CUG GAG AUU) predicted to bind to at least one of the proteins comprising the translation complex eukaryotic Initiation Factor 3 (eIF3; [11–14]). Published reports identify the p120 subunit of eIF3 as the one binding to the apical loop of the domain III of HCV 5'UTR, and p170 (also called p160) as binding the stem portion of the domain III. The oligonucleotide used for these studies contains the apical portion and part of the stem portion of domain III and, therefore, might bind one or both of these subunits. Interestingly, the binding of p170 (p160) to the stem of domain III is position-independent (i.e., not constrained by other structural elements on the mRNA [12, 13]).
Sequence analysis of surrogate peptide binders to HCV using MEME (Motif Elicitation Program) and other peptide sequence alignment programs identified a consensus sequence TxRLL found in four peptides binding to the HCV mRNA (Figure 5). This motif was unique to HCV and not found in peptides derived from any of the other mRNA pans. Interestingly, the TxRLL-containing surrogates were found by two investigators in the laboratory, using the 20 mer or 40 mer random libraries under different experimental conditions. Database searches using Patternfind at the ISREC server were performed using parameters appropriate for short protein queries and were successful in identifying a human gene product, subunit p170 of eIF3 (Figure 3). Searches using the consensus region as the query likewise identified sequence homology with the large subunit p170 of eIF3. These results fit our partner criteria 1–3 (Table 1). Interestingly, other amino acids in these surrogates were found to be identical to residues flanking the TxRLL motif in p170 on both the amino and carboxy sides (Figure 5). Thus, the library contained peptide surrogates binding to the HCV target mRNA as well as containing sequence information identifying the natural interacting partner and predicting the putative contact amino acids on the p170 subunit of eIF3. The fact that multiple surrogates had the TxRLL motif suggests that this region of p170 is necessary and critical for binding of HCV mRNA to eIF3.
Alignment of the TNFβ surrogate peptide KcB7 with its cognate receptor TNFR1 (p55). Peptide KcB7 was isolated by panning TNFβ. The peptide has the sequence RKEMGGGGGPGWSENLFQ and a BLASTp search revealed identity to amino acids 77–81 (RKEMG) and amino acids 107–113 (WSENLFQ) on TNFR1 (p55). RKEMG is involved in binding of TNFR1 to the A subunit of TNFβ while WSENL binds to several amino acids in the C subunit of TNFβ
Panning of TNF-β
In a separate study, we panned the immune cytokine TNF-β (lymphotoxin α, Lt α; [15–17]) using a highly stringent protocol involving both positive (vs. TNF-β) and negative (vs. TNF-α, TNFR1 and TNFR2) selection. One of these peptides, designated KcB7, had the amino acid sequence RKEMGGGGGPGWSENLFQ. A Blastp search, using several different queries revealed TNFR1, which is the natural biological partner of TNFβ (Figure 5). Interestingly, the other cognate partner (i.e., receptor TNFR2) was not identified. Closer examination of the complementary sequences revealed that the short N-terminal sequence RKEMG and the C-terminal sequence WSENLFQ were identical to regions on TNFR1 (amino acids 77–81 and 107–113 respectively). Although not complete, these segments corresponded to amino acids within two critical ligand:receptor contact domains [18]. In the case of the N-terminal grouping, the surrogate contained 5 of the 15 amino acids of the 77–81 contact domain whereas in the C-terminal grouping, the surrogate contained 6 of the 9 amino acids identified within the 107–113 contact domain. It is also worth noting that, on the receptor, these two domains are not contiguous but are separated by 25 amino acids whereas only a 6 amino acid hydrophobic stretch separates the two noncontiguous contact-mimicking domains in the surrogate. Therefore, the seven amino acid linker may provide the appropriate molecular distances needed for TNFβ binding to the receptor.
Robustness of the System
The successful isolation of a useful surrogate may seem an improbable task especially in cases where nothing is known about the hotspot surface(s) on a target. In reality, biopanning using the RAPIDLIB® library has allowed, (in >90% of cases) the identification of partner-specific peptides among the >1010-11independent clones that are expressed within this library [[7, 19, 20] and unpublished data]. More often than not, between 10 and 100 different surrogates have been found for any one target panned and >75% of the targets gave rise to surrogates that bound to regulatory hotspots on the target [[7] and unpublished data]. Several important facts are critical to the success of the process. In the first place, there must be a high degree of diversity within the library so that one surrogate can be found for each protein target. Additionally, It appears that panning seems to favor the enrichment of regulatory surrogates versus irrelevant peptides. In our experience, the large number of random peptides includes a broad sampling of linear and conformational protein surfaces sufficient to present at least one low affinity "complement' for any target's surface [19, 20]. The immune system's antibody repertoire arises through a process [21–24] similar to in some aspects to what occurs during the panning process. Initially, immunoglobulins are synthesized containing multiple small contact domains (i.e., CDR or Complementarity Defining Regions) to provide a "rough" complement of an antigen's surface. Subsequently, antibody/antigen binding is improved via mutational events within the antibody genes to produce a complement with higher affinity and selectivity. Biopanning works in an analogous manner by initially enriching for peptides with low affinity for a specific target. However, surrogates may only need to contain some amino acids identical to those on the partner to bind with sufficient "avidity" rather than affinity.
Other methods that have been used extensively for high throughput partner identification include the yeast two-hybrid system [25]. While the two hybrid approach is popular, it has a number of inherent problems including a high potential for false positives, the inability to use non-protein targets such as mRNA or membrane bound/extracellular proteins and the inability to address postranslational modifications on a target. The generation of surrogates, on the other hands, is target independent and their utility for partner identification resides in the computational analysis of the peptide's sequence. Since our libraries contain totally random peptides ranging from 20 and up to 40 amino acids in length, there are no known restrictions on the amino acids that can be selected to create the surrogate's 'complementary' surface [7]. Thus, the examples described in this report relate to the utility of the surrogate approach for finding the cognate receptor for both protein and non-protein targets. In the case of the surrogates for both HCV-mRNA and TNFβ, it is clear that the large diversity and size of the original library was, in fact, critical to their successful isolation since libraries of <20 amino acids would not have contained either the KcB7 peptide or the HCV-specific surrogates.
In addition to the data presented in this paper, we have screened other targets using this approach. While the expected natural partners were found for many of the proteins, there were instances where surrogates were generated but lacked partner information (e.g., IGF-1R, growth hormone receptor, insulin receptor [[7] and manuscripts in preparation]). There are several possible explanations for these results. While our libraries are large and diverse, it is probable that identification of a surrogate peptide with partner information is a rare event. With that in mind, it may require the isolation and sequencing of large numbers of clones (perhaps >500/target) in order to find the appropriate surrogate for partner identification. On the other hand, some targets may have complex or unusual protein:protein contact sites that preclude generation of a surrogate with partner information. Ongoing experiments will address both of these possibilities. Surrogates have also been found to have the minimal structural content necessary to induce a pharmacological effect on any target in addition to their use in partner identification. Most of our surrogates have been shown to have either agonist or antagonist activity in the appropriate biochemical and/or biological models ([7]manuscripts in preparation). The surrogates were also able to subdivide large contact surfaces into smaller contact domains through which target activity can be modified [7]. These attributes point to surrogate use in phenotyping and validating novel genes whose functions are unknown and for which no known partners exist. Finally, it is worthwhile to note that surrogates have been used to develop competitive Site Directed Assays (SDAs) for essential sub-domains thereby allowing their use in high throughput screening of large combinatorial libraries of small molecules (unpublished data). In our experience, almost all of the peptide surrogates isolated from these complex libraries by our various panning procedures bind to regulatory hot spots on varied targets. This non-random association between a surrogate and a target's pharmacologically active site assures a high degree of probability that, once found, surrogates will have utility for the rapid development of SDAs capable of identifying small molecules of pharmacological importance.
The results of these experiments support the use of in vitro panning procedures with our highly complex and random 20–40 mer peptide libraries as a method to obtain information on large numbers of protein partners and enable the elucidation of biologically relevant protein networks. This post-genomic approach can be automated to increase the number of known and unknown genes and gene products that can be used as targets for partner identification as the first step in the drug discovery process. Additionally, the surrogates isolated in such studies would be prime candidates for phenotyping and target validation through their ability to regulate target activity and for identifying small molecule drug leads through their use in Site Directed Assays. Overall, the surrogates can be seen as providing a discovery continuum by bridging the gap between functional genomics and proteomics and modern drug discovery.
Oligonucleotides for the mRNA pans were synthesized by Dharmacon Research (Lafayette, CO) and used for the experiments after heat treatment and re-annealing (65° for 10' and slow cooling at room temperature). The APP and IGF oligo sequences are proprietary to Message Pharmaceuticals. The following sequence was used to obtain the HCV surrogates: 5'-biotin'AA UUG CCA GGA CGA CCG GGU CCU UUC UUG GAU CAA CCC GCU CAA UGC CUG GAG AUU-3'. The sequence for RRE (Rev-response element) has been previously published [10]. Streptavidin coated plates were obtained from Pierce (Rockford, IL). TNFR1, TNFR2, TNF-α and TNF-β were obtained from R&D Systems (Minneapolis, MN) and reconstituted according to the manufacturer's instructions. The E. coli, strain TG1 (genotype = K12Δ(lac-pro), supE, thi, hsdΔ5/F' [traD36, proAB, lacIq, lacZΔM15]) was obtained from Pharmacia (Piscataway NJ).
Design of the Primary Peptide Libraries
DNA fragments coding for peptides containing 40 random amino acids were generated by a PCR approach using synthetic oligonucleotides as previously described [7, 26]. Peptides are expressed on the capsid protein pIII of the phage at low copy number (1–2 peptides/phage).
Construction of secondary cell libraries
Amino acid mutations were introduced at the oligonucleotide level under controlled conditions [7]. This oligonucleotide was used as the template in a PCR reaction with two shorter 5' biotinylated oligonucleotide primers contributing the restriction sites. The library was then produced essentially as previously described (above). Cell transformants were pooled and an aliquot was plated to determine the total number of transformants. The diversity of the secondary cell libraries >1010 independent clones per library.
Biopanning and ELISA protocols
For the TNF experiments, 6 wells of a 96-well microtiter plate (Maxisorb Immuno plates, Nunc) are coated with target at concentrations ranging from 5–50 ng/well and are incubated overnight at 4°C. At the same time, an aliquot of E. coli TG1 cells is inoculated into 2x YT media and grown overnight at 37°C. The next day, unbound antigen is removed and the coated wells are blocked with 300 μl of 2% non-fat milk in PBS (NFM-PBS) for one hour at room temperature. The phage libraries (in NFM-PBS) are thawed and 100 μl of each library is added to the antigen-coated wells and the plates are incubated for 3 hours at room temperature. Each well is washed 13 times with PBS and the phage eluted with 100 μl of 50 mM glycine-HCl containing 0.1% BSA (pH2.2) following a five minute incubation. The eluted phage are pooled, neutralized with 100 μl of 1 M Tris-HCl (pH 8.0). The eluted phage were then panned vs. other members of the TNF family as a way to obtain peptides specific to the original target (Subtractive panning). For example, phage, initially panned vs. TNF-α would be subsequently panned vs. TNF-β, TNFR1 and TNFR2 to remove peptides cross-reacting with the other members. After the final panning, the phage are added to 10 ml of log phase E coli TG1 (OD600 = 1.0), and amplified in 2x YT-glucose medium for one hour at 37°C. Helper phage (M13K07) and ampicillin are then added and the cells are incubated for an additional hour at 37°C. The cells are pelleted at 3500 RPM for 10 minutes, resuspended in 2x YT-AK medium (YT medium containing ampicillin and kanamycin) and incubated overnight at 37°C. The next day, the infected bacterial cells are centrifuged at 3500 RPM at 4°C for 10 minutes and the pellet discarded. The supernatant contains the phage and is precipitated with ¼ volume of 20% PEG-8000 in 1.6 M NaCl by incubating on ice for 1 hour. The precipitant is centrifuged at 10,000 RPM at 4°C for 10 minutes and the precipitated phage pellet resupended in about 1 ml of NFM-PBS. The phage is then used for the next round of panning. Normally, 3–4 rounds of panning are done for secondary libraries. Usually, 96 random clones are picked from rounds 3 and 4 and grown in 96 well cluster plates as a master stock.
For the mRNA pans, all solutions and surfaces are pretreated with DEPC or RNaseZap (Ambion, Austin TX), respectively, to eliminate RNase contamination that may compromise the integrity of the RNA. The biotinylated – RNA target diluted to 1 mg/ml in binding buffer (PBS containing 1 mM MgCl2), denatured at 65°C for 5 min and reannealed by slow cooling to room temperature. Re-annealed mRNAs are stored in small aliquots (10–25 μl/tube) at -20°C. Microtiter wells are treated with RNaseZap (Ambion) before use. One hundred microliters of RNA solution diluted to 2.5 ng/ μl was added to an appropriate number of wells in a 96-well microtiter plate precoated with Streptavidin (Pierce) and incubated for 1 hr at room temperature. Unbound streptavidin is then blocked with 50 μl of 2 mM biotin at room temperature for 1 hr. Panning, with slight modifications, proceeded as described for TNF-β.
For phage rescue prior to ELISA analysis, 40 μl of master stock is transferred from each master to another set of cluster tubes containing 400 μl of 2x YT-AG (ampicillin and glucose) and helper phage (final concentration of 5 × 1010/ml). The tubes are incubated at 37°C with constant shaking for two hours. The cultures are centrifuged at 2500 × g at 4°C for 20 minutes, the supernatant is discarded, and the bacterial pellet is resuspended in 400 μl of 2x YT-AK (ampicillin and kanamycin) medium and is incubated overnight at 37°C. At that time, the cells are removed by centrifugation at 2500 × g and the supernatants used for ELISAs (see below).
Enzyme Linked Immunosorbent Assays (ELISA)
For the TNF-β surrogate ELISAs, each well of a MaxiSorp Immuno plate (Nunc) is coated with 50 μl of target (1 μg/ml) overnight at 4°C. In all cases, the wells are blocked with NFM-PBS for 1 hour at room temperature. Phage is added at 100 μl/well and the plates incubated for 2–3 hours at room temperature. After washing 3x with PBS-Tween (DPTC-treated PBS for the mRNA), plates are probed with an anti-M13 antibody conjugated to horseradish peroxidase (1:3000 in PBS-NFM) for 1 hour at room temperature followed by addition of 100 μl of ABTS for 15–30 minutes at room temperature. The OD is measured using a SpectraMax Microplate Spectrophotometer (Molecular Devices) at 405 nm after a 30 minute incubation at room temperature.
The mRNA ELISAs are performed in a similar manner except streptavidin-coated microtiter plates are blocked with PBS containing 2% non fat milk for 1 hr at room temperature, treated with RNaseZap, then coated with biotinylated RNA target (100 ng/well) by incubation for 1 hr at room temperature. Superasin was added to the wells prior to addition of 100 μl/well of phage from isolated clones and incubated at room temperature for 2 hr.
Steps in computational analysis to identify natural partner
Once a surrogate peptide binder has been identified and shown to bind specifically to its target, it is subjected to partner analysis using several different database search programs. In the initial step, the entire peptide sequence and consensus motifs (if found) are entered into an Advanced BLAST search http://www.ncbi.nlm.nih.gov/blast/blast.cgi?Jform=1: using the following parameters:
i. Programs: blastp, tblastn
ii. Databases: protein and nucleotide databases including dbest (ESTs), dsts (STSs) and htgs (unfinished high throughput genomic sequences)
iii. Expect value: 1000 to 20000
iv. Matrix: PAM30 or PAM70
v. Query: Consensus motif alone and varying combinations of sequence at the N- and C-terminal ends
In subsequent steps, motifs identified by sequence alignment programs like MEME (Multiple EM for Motif Elicitation), http://meme.sdsc.edu/meme/website/intro.html can also be used to search the available databases using MAST (Motif Alignment and Search Tool, http://meme.sdsc.edu/mem/website/mast-intro.html). Motifs and consensus domains can also be used as query patterns to search the protein databases using Patternfind http://www.isrec.isb-sib.ch/software/PATFND_form.html.
For Patternfind, the following parameters are used:
i. Databases: Nonredundant, Swissprot, TREST and TRGEN
ii. Limit: Between 10 and 5000
iii. Query: Consensus motif alone and varying combinations of sequence at N- and C-terminal ends
Data obtained from the various searches are analyzed under the following conditions:
i. Results of different searches should be analyzed independently and then together to look for similar classes of proteins (e.g., Nucleic acid binding proteins, kinases) that may emerge.
ii. Pick some of the best matches that show up in more than one kind of search (e.g. Same protein/ORF picked up by BLAST searches using different parameters, or by both BLAST and Patternfind) and compare sequence of protein in this region with other peptide surrogates containing this motif.
iii. Look at potential significance of protein interaction in the context of the cellular function of target
The criteria for a partner hit are listed in Table 1. Unless there is an exact match (Criterion No. 1), the potential hit has to match at least two of the criteria described in the table to be considered a partner hit.
Drews J: A historical perspective. Science. 2000, 297: 1960-1967. 10.1126/science.287.5460.1960.
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Gallop MP, Barrett RW, Dower WJ, Fodor SP, Gordon EM: Applications of combinatorial technologies to drug discovery. I. Background and peptide combinatorial libraries. J Med Chem. 1994, 37: 1233-1251.
Yanofsky SD, Baldwin DN, Butler JH, Holden FR, Jacobs JW, Balasubramanian P, Chinn JP, Cwirla SE, Peters-Bhatt E, Whitehorn EA, Tate EH, Akeson A, Bowlin TL, Dower WJ, Barrett RW: High affinity type I interleukin 1 receptor antagonists discovered by screening recombinant peptide libraries. Proc Natl Acad Sci USA. 1996, 93: 7381-73. 10.1073/pnas.93.14.7381.
Wrighton NC, Farrell FX, Chang R, Kashyap AK, Barbone FP, Mulcahy LS, Johnson DL, Barrett RW, Jolliffe LK, Dower WJ: Small peptides as potent mimetics of the protein hormone erythropoietin. Science. 1996, 273: 458-463.
Cwirla SE, Balasubramanian P, Duffin DJ, Wagstrom CR, Gates CM, Singer SC, Davis AM, Tansik RL, Matteakis LC, Boytos CM, Schatz PJ, Baccanari DP, Wrighton NC, Barrett RW, Dower WJ: Peptide agonist of the thrombopoietin receptor as potent as the natural cytokine. Science. 1997, 276: 1696-1699. 10.1126/science.276.5319.1696.
Blume AJ, Beasley J, Goldstein NI: The use of peptides in Diogenesis: A novel approach to drug discovery and phenomics. Biopolymers. 2000, 55: 347-356. 10.1002/1097-0282(2000)55:4<347::AID-BIP1007>3.0.CO;2-J.
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Zemmel RW, Kelley AC, Karn J, Butler PJG: Flexible regions of RNA structure facilitate co-operative Rev assembly on the Rev-response element. J Mol Biol. 1996, 258: 763-777. 10.1006/jmbi.1996.0285.
Buratti E, Tisminetzky S, Zotti M, Baralle FE: Functional analysis of the interactions between HCV 5' UTR and putative subunits of eukaryotic translation initiation factor eIF3. Nuc Acid Res. 1998, 26: 3179-3187. 10.1093/nar/26.13.3179.
Odreman-Macchioli FE, Tisiminetzky SG, Zotti M, Baralle FE, Buratti E: Influence of correct secondary and tertiary RNA folding on the binding of cellular factors to the HCV IRES. Nuc Acids Res. 2000, 28: 875-885. 10.1093/nar/28.4.875.
Kruger M, Berger C, Li QX, Welch PJ, Trinz R, Leavitt M, Barber JR, Wong-Staal F: Identification of eIF2Bγ and eIF2γ as cofactors of hepatitis C virus internal ribosome entry site-mediated translation using a functional genomics approach. Proc Natl Acad Sci USA. 2000, 97: 8566-8571. 10.1073/pnas.97.15.8566.
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Ravera MW, Carcamo J, Brissette R, Alam-Moghe A, Dedova O, Cheng W, Hsiao KC, Shen KI, Tang P, Blume A, Mandecki W: Identification of an allosteric binding site on the transcription factor p53 using a phage-displayed peptide library. Oncogene. 1998, 16: 1993-1999. 10.1038/sj.onc.1201717.
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The authors would like to acknowledge the expert technical assistance of, Darek Galkowski, Kwok-Lam K. Ho, Leigh-Ann Miloscia, Adrienne Parratt and Huong Vuong.
DGI BioTechnologies, Inc., Edison, NJ, USA
Renuka C Pillutla
, Ku-chuan Hsiao
, Renee Brissette
, Paul W Fletcher
, Michael Lennick
, Arthur J Blume
& Neil I Goldstein
Message Pharmaceuticals, Inc., Malvern, USA
Paul S Eder
& Tony Giordano
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Correspondence to Neil I Goldstein.
Renuka C Pillutla, Ku-chuan Hsiao contributed equally to this work.
Random Peptide
Contact Domain
Natural Partner
Modern Drug Discovery
Surrogate Peptide

redpajama_set_name: RedPajamaCommonCrawl


The Horace Greeley House was the country home of the Greeley family from 1864 until 1873. It is one of three homes that the Greeleys owned in Chappaqua. Mr. Greeley was the editor and publisher of the New York Tribune, and most of the time he lived in New York City.
The Main Floor consists of three primary rooms. The Front Parlor was used for formal entertaining and receiving guests. Over the handsome fireplace, there is a print which shows the Greeley family in 1872, the year that Greeley ran for president. The Dining Room, painted in a bright shade of red, overlooked Greeley's Farm. On one wall a print shows Greeley's campaign slogan, "Yours for Universal amnesty and impartial suffrage." The Music Room is one of the rooms in the portion added by the Greeleys. Music was important to the Greeley family. This room includes a box pianoforte similar to the one belonging to the Greeleys. One portion of the current Research Room served at the Butler's Pantry.
The Upper Floor also consists of three primary historic rooms. When Greeley enlarged the house, he altered and modernized the second floor, transforming it from Federal to Victorian style. He raised the ceilings and put in tall windows, including the French doors leading to the balcony. The Family Parlor was a more private sitting room than the downstairs parlor. In this room are displayed many items connected to Greeley's professional career and political activities. The Girls' Bedroom, shared by Gabrielle and Ida, is a bright and airy room, opening out onto the balcony through the three French doors. Today we use this room for special exhibitions. The Master Bedroom is where Mary Greeley spent much of her time. The only photograph we have of her hangs on the wall by the door to the stairway. As in the dining room, the back wall is where the original house ended, and it contained two windows overlooking the farm. The Maid's Room is now for use by the staff.
The Meeting Room behind the bedroom is part of the new section of the house and is a large room used for meetings and special exhibitions. This section of the house was added in the 1950s when the house was used as a gift shop.
The Ground Floor consists of a Kitchen and Laundry Room. In the Kitchen we show several utilitarian objects such as a spinning wheel and yarn winder which belonged to Greeley's mother, and many other objects that one would find in a Victorian kitchen. The Laundry Room is currently used for storage.
The grounds, which are handicapped accessible, contain period Perennial and Herb Gardens which feature unusual species and plants once used for medicinal purposes.


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Tuesday, July 27, 2021 1:00 pm (CDT) The Tao Te Ching and the Divine Feminine
Author, professor, and Episcopal priest Rosemarie Anderson, Ph.D., talks about her new translation of and commentary on Lao Tzu's great…
Tuesday, July 13, 2021 1:00 pm (CDT) Opening to Grief with Claire B. Willis and Marnie Crawford Samuelson
The authors of Opening to Grief: Finding Your Way from Loss to Peace join the show—social worker, lay Buddhist chaplain, and author…
Tuesday, July 6, 2021 1:00 pm (CDT) Loving and Living Your Way Through Grief
Learn how to reclaim and activate joy and caring in the face of loss with author and speaker Emily Thiroux Threatt.
Tuesday, June 29, 2021 1:00 pm (CDT) Chakra Healing Therapy with Glen Park
Discover how to awaken spiritual energies and heal emotional wounds with teacher and healer Glen Park.
Tuesday, June 22, 2021 1:00 pm (CDT) When It's Time to Say Goodbye to a Beloved Pet
Get help in preparing for the transition of your beloved pet with author and animal bereavement specialist Angela Garner.
Tuesday, June 15, 2021 1:00 pm (CDT) Transcendent Leadership with Richard Anderson
Author, coach, and spiritual teacher Richard Anderson discusses the eight dimensions of awakened leadership to help live a more…
Tuesday, June 8, 2021 1:00 pm (CDT) A Story of Karma with Michael Schauch
Find love and truth in the lost valley of the Himalayas with Michael Schauch, whose journey transformed his life. Michael shares the…
Tuesday, June 1, 2021 1:00 pm (CDT) Vibrational Sound Healing with Erica Longdon
Metaphysician, radio host, writer, and healer Erica Longdon shares her knowledge of sonic vitamins using tuning…
Tuesday, May 25, 2021 1:00 pm (CDT) Where Wonder Lives with Fabiana Fondevila
Explore practices for cultivating the sacred in your daily life with author, storyteller, and teacher Fabiana…
Tuesday, May 18, 2021 1:00 pm (CDT) Blue-Collar Spirituality with Michael McMorrow
Paul discusses finding a God that works with Religious Science minister Michael McMorrow.


There's hard work and then there's Hardy work.
What is it about celebs and gymnastics we like? Maybe just the unusual nature of the photos, the way they depart from typical promo portraits. So today, joining previous bendy celebs such as Danielle Darrieux, Joey Heatherton, and Constance Dowling is French actress Sophie Hardy—yes, again. When we posted her as a femme fatale last month we hadn't seen this photo yet. It had to be shared. You may also remember Hardy recently fronting a beautiful Japanese poster for The Trygon Factor. This image comes from around the same time as the movie, circa 1968. And the shot below shows her right side up, possibly lightheaded, but none the worse for wear.
The ongoing process of discovery that motivates us to maintain this website continues. We'd never heard of French actress Sophie Hardy before watching her in The Trygon Factor, a movie we talked about a couple of months ago. Since then we've found her in numerous eye-catching promo photos. Which means you'll be seeing her again. This winner comes from around 1965.
If at first you aren't impressed try Trygon again.
You may have noticed we're really digging into our collection of Japanese posters lately. Here's another. It was made for Das Geheimnis der weißen Nonne, a West German-British movie based on Edgar Wallace's novel Kate Plus Ten and released in English as The Trygon Factor. Basically, a Scotland Yard investigator investigates a series of robberies and finds that a respectable English family and group of nuns living in a country manor together are not what they seem. Stuart Granger and Susan Hampshire star, but the star of the poster is Sophie Hardy, which just goes to show the Japanese distributors knew exactly how best to promote the film. We saw the movie several years back and weren't impressed, but we watched it again last night and this time found it pretty entertaining for the type of goofy crime thriller it is. We make no guarantee, though. Das Geheimnis der weißen Nonne-The Trygon Factor premiered in Japan as Dai gôtô-dan today in 1968.


Different people find different ways to say thank you, and this wonderful lady is saying thank you to our rescue by publishing a book you can purchase and keep track of all your amaizng events you share with your sugar gliders. Thank you Leslie! This would also make a wonderful gift for the sugar glider "parent" in your life!
A safe, appropriate sized, exercise wheel that is truly silent.
An easy to set up safe place for bonding and play time with your gliders.
Visit Molly's Playhouse on Facebook, or contact directly for a variety of handmade items, including the City Pouch bonding pouch!
Visit Lovely Sugar Stitches on Facebook or contact directly for a variety of handmade toys and cage sets.
There are many sources out there for buying sleeping pouches, cage sets, bonding pouches, toys and other fun accessories for your sugar gliders. Here are a couple of our favorite vendors! Be sure to always monitor your items for safety. Loose threads, unraveling fabric, holes chewed in fabric or tabs, can all be unsafe in a sugar glider's environment.
Check out Sugar Glider Joy! They sell wonderful, glider safe cage sets, accessories, bonding pouches, and the original Cozy Clip set. Their goal is glider safe products in every glider home!
Spike's Ultra Premium Hedgehog food is designed specifically for pet hedgehogs.
The Midwest Guinea Habitat system with the optional lid, is a good size for a pygmy hedgehog!
The Carolina Storm Bucket Wheel is the best hedgehog safe wheel we have tried!
Different people find different ways to say thank you, and this wonderful lady is saying thank you to our rescue by publishing a book you can purchase and keep track of al your amazing events you share with your hedgehog! Thank you, Leslie! This would also make a wonderful gift for the hedgehog "parent" in your life!


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One of the world's thickest mountain glaciers is melting because of global warming
Doyle Rice
Thick mountain glaciers are melting in Alaska as temperatures warm.
One of the world's thickest mountain glaciers is finally succumbing to global warming, a new analysis reports.
The Taku Glacier north of Juneau, Alaska, has started to retreat as temperatures rise, said Mauri Pelto, a glaciologist at Nichols College in Massachusetts.
Up until now, of the 250 glaciers he has studied, all had retreated except one: Taku Glacier. But an analysis shows Taku has lost mass and joined the rest of the retreating glaciers.
"This is a big deal for me because I had this one glacier I could hold on to," Pelto told NASA's Earth Observatory. "But not anymore. This makes the score climate change: 250, and alpine glaciers: 0."
Taku is one of the thickest known alpine glaciers in the world, measuring 4,860 feet from surface to bed.
A study about the glacier's retreat was published last month in the journal Remote Sensing.
Overall, global warming continues to melt away our planet's glaciers, which lose up to 390 billion tons of ice and snow every year, a study published earlier this year suggests.
Glacier and Ice Sheet Melt


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Lali Khalid Photography Exhibit at Handwerker Gallery
By Mickie Quinn, August 26, 2021
First Light, the Skylarks Sing - work by Lali Khalid
Opening Reception: Thursday, September 2, 5-7 p.m.
Handwerker Gallery - Gannett Center
Artist Talk: Thursday, September 16, 6 p.m.
Artist book launch: Thursday, October 7, 5:30 p.m.
'Home. In my heart, beating far away.' https://www.artvoicesartbooks.com/shop/lalikahlid with introduction by Raza Rumi, Director (IC Park Center for Independent Media).
Mehreen (Lali) Khalid came to the United States from Pakistan in 2007 as a Fulbright Scholar and attended Pratt Institute, NY where she received an MFA in Photography in 2009. After returning to Pakistan for two years, she immigrated to the United States in 2011.
Issues relating to diaspora, identity, family and home dominate Khalid's photographic inquiry. Her images focus on cultural and private conflicts, broaching these topics obliquely through the emotive effects of natural light, and subdued narrative allusion. By photographing primarily in color, and using the startling qualities of "found light," Khalid's photographs exist between a mood of quiet longing.
Lali Khalid has shown her work in numerous exhibitions throughout Europe, Pakistan, and the U.S., including most recently at Light Work, Roy G Biv and Devos Museum. She is currently an Assistant Professor of Photography in the Roy H. School of Communications, Ithaca College, Ithaca, NY.
"First Light, the Skylarks Sing" exhibit featuring works by Lali Khalid at the Handwerker Gallery on September 2nd through October 13, 2021.
Individuals with disabilities requiring accommodations should contact Mara Baldwin at mbaldwin@ithaca.edu or 607.274.3548. We ask that requests for accommodations be made as soon as possible.


After four and a half hours of journey on the Jaipur Express, I arrived in Jaipur. As it happens when visiting a new place, the first few hours in the city can be a little daunting. Especially for those without previous experience traveling around India, as it was my case. Temperatures can easily reach 35ºC or even 40ºC before noon, and the roads can be a little intimidating with all types of vehicles – including buses, cars, motorcycles, rickshaws, bicycles, elephants, and even camels – traveling in all directions. It only takes a little bit before starting to immerse in the charm of Jaipur and to start to discover everything that the city has to offer.
The old city center is encircled by a wall and a number of beautiful gates. These gates, along with a number of buildings situated within and around the old city center, display a distinct pink colour that give Jaipur the name of the 'pink city'. Visiting the old city center is a delight for all senses: the colourful bazaars, the amazing architecture, delicious street food all around, and the beautiful traditional clothing that a large number of people continue to wear.
Eating in Jaipur is also a treat. I had the pleasure of trying homemade food, prepared by the welcoming and warm family that hosted me during my stay in Jaipur; street chai and tasteful snacks from a number of street vendors; and traditional and modern Indian food from amazing restaurants that would easily be in the top ten restaurants that I have visited across the globe. I especially recommended Tapri and the terrace of the Grand Peacock Restaurant. Also, in case you are interested in high-quality home accommodation in Jaipur, I strongly recommend checking-out Shan's place. Shan and their family are amazing hosts and their house is beautifully decorated.
About 10 km away from Jaipur, lies the small town of Khania-Balaji. The town is home to the pilgrimage site called Galta Ji, commonly known as 'Monkey temple'. The route to the site through the small village is really beautiful, and the architecture and sights from the top of the temple are just breathtaking. Not to mention, the fun that it is to watch the playful monkeys that abound around the temple.
A short ride away from Jaipur, lies the magnificent Amer Fort, one of the six Jaipur forts declared as World Heritage Site by UNESCO. The location, dimension and architectural beauty of this site, alone, make it worth a visit to Jaipur. The only regret that I have about this trip, is not having had more days to visit the other forts and the many other sites spread across the city.
Having had the opportunity to visit many of the main cities and capitals in Europe, I never thought that I could be that surprised by another European city. I was wrong. Lisbon is just amazing. Travelling to a new destination normally involves a couple of main attractions that make the trip worth it, and then, a few secondary elements that round up the experience. In some places, the main attraction is the seaside; in others the food; in others the street life; in others a historic building or monument; in some place the weather; in some place the people; and so on.
Well, this is not the case with Lisbon. Lisbon has everything and has it in abundance. Some places in Lisbon, such as Praça do Comercio, Castelo de Saõ Jorge, Mosteiro des Jeronimos, Parque Eduardo VII or Ponte 25 de Abril could easily be the main attraction in any other destination; and any of those sites, alone, would attract millions of visitors every year.
Lisbon has them all and that is just a fraction of what the city has to offer. The city, sprawled across seven hills alongside the Tagus river, also offers a magical experience when wandering around the city. Chiado, Barrio Alto, Baixa and Alfama, are only a few of the many picturesque neighbourhoods that bring together centuries of history; uncountable parks and plazas; walls full of colours and beautiful azulejos (tiles); the famous Portuguese pavement that make up the unique patterns of streets and plazas; and a number of charming means of transport, including tram, tuk-tuks, lifts and funiculars.
As if this was not enough, Lisbon also has an outstanding culinary offer based on a mediterranean diet and a vast array of fresh and high-quality ingredients, including sea food, vegetables, fruits and juices, cheese, olives, cold cuts, wine and extraordinary bread.
Finally it is also worth mentioning the friendliness and warmth of Portuguese people. Everyone, from the officers that received us at the airport, to the taxi and bus drivers, to the waiters and musicians, to our Airbnb host; every Portuguese we met was always smiling and always willing to help.
About six kilometres away from the city center, lies the beautiful parish of Belém. A trip to this part of Lisbon is a must in order to visit the Mosterio des Jeronimos and the Belem tower. The route is quite scenic with Ponte 25 de Abril and Cristo Rei as main landmarks. Once in Belém, it is a must to try the Pastéis de Belém that have been in business for almost 180 years.
Less than an hour away from Lisbon, lies the beautiful town of Cascais. Located in the estuary of the Tagus river, Cascais provides a wonderful opportunity to enjoy the sunset and to sight the beautiful beaches and the colourful Farol de Santa Marta.
In September 2013, I had the chance to visit Beijing, in China. Having lived in Mexico City for over 30 years, I love big cities. Beijing had a special vibe combining old traditional architecture and history with the buzz of a modern and growing city. Not to mention the amazing food and people.
One hundred and thirty kilometers north from Mexico City, lies the beautiful town of Huasca in the state of Hidalgo. Established as a mining town, Huasca is home to some of the finest and most amazing 'Haciendas' (estates) in the central part of Mexico. Huasca also is surrounded by enchanting natural spots, including the Basaltic Prisms, amazing geometric rocks carved by water over millions of years.


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January 7, 2019 by NOW National
Celebrating The Life and Work of Bernice Sandler
Statement by NOW President Toni Van Pelt:
Bernice Sandler, who passed away January 5 at her home in Washington, D.C., was widely known as the "Godmother of Title IX." After earning a doctorate at the University of Maryland in 1969, she applied for one of seven teaching positions in her department but was told that she wouldn't be considered for any of them.
"Let's face it," a male colleague told her. "You come on too strong for a woman." When she applied for another position, the researcher in charge of hiring said he didn't hire women because they stayed home too often to care for sick children. Later, an employment agency dismissed her as "just a housewife who went back to school."
Bernice Sandler then embarked on a lifelong mission to change the culture of sex discrimination on college campuses. She spent decades documenting, investigating and working to change the arbitrary limits and sexist standards that held women back academically and professionally. Her work led to the passage in 1972 of Title IX, the landmark legislation that banned sex discrimination in federally funded educational institutions.
Bernice Sandler was inducted into the National Women's Hall of Fame in 2013 when she was lauded for her efforts that led to the first federal investigation of sex discrimination on campuses.
Bernice Sandler was an inspiration to countless women who were inspired by her example and empowered by her accomplishments. She will be missed but never forgotten.
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You are at:Home»News»Arts»'It is important that all parties pull together to ensure our vibrant arts, creative businesses and creative institutions are not impacted'
'It is important that all parties pull together to ensure our vibrant arts, creative businesses and creative institutions are not impacted'
By Maria Roberts on	 June 24, 2016 &nbspAnalysis	Arts, News
Early statements from the UK arts sector today (24 June) on the Referendum result that will see the UK begin the process to leave the EU
'Our country is a creative powerhouse and we know for sure that the creative economy will continue to be at the centre of our economic future'
Creative England
'The UK is full of great businesses fuelled by enormous creativity and the free movement of ideas, goods and talent. Our country is a creative powerhouse and we know for sure that the creative economy will continue to be at the centre of our economic future; maintaining our competitive edge in the light of increasing global competition is vital. It is important that all parties pull together to ensure our vibrant arts, creative businesses and creative institutions are not impacted negatively by the uncertainty and turmoil of the next few weeks and months. And it is vital we build sustainability for our long-term future rather than short term expediencies.
'But creativity is not only a driver of our economic prosperity: it's also at the heart of how we express ourselves and pull our communities together. As the UK looks to build a new relationship with Europe and the rest of the world safeguarding the institutions, funding and ecosystems that support creative talent and creative businesses to thrive will be of huge importance.' – Caroline Norbury MBE, CEO of Creative England.
'The ABO's next step is to build on our existing good relations with the government and engage positively with whichever Ministers take responsibility from here on, to ensure the best possible outcome for our members'
Association of British Orchestras (ABO)
Following the Referendum decision to leave the EU, the ABO is deeply concerned at the potential impact on its members.
The prospects for the nation's public finances are worrying, and may affect the implementation of Orchestra Tax Relief, which has not as yet received Royal Assent, and lead to further reductions in public funding for the arts and local authorities.
We will need the new leadership of this country to give us guarantees as to continued freedom of movement across Europe's borders for our orchestras, artists and orchestral musicians, and whether the many pan-European regulations that currently affect our sector, from VAT Cultural Exemption to harmonisation of radio spectrum, Noise at Work to the Digital Single Market, will still apply.
The worst outcome for our members will be additional uncertainty, bureaucracy and expense, allied to a worsening of their financial viability. The ABO's next step is to build on our existing good relations with the government and engage positively with whichever Ministers take responsibility from here on, to ensure the best possible outcome for our members – Mark Pemberton, director, Association of British Orchestras
International Artist Managers's Association (IAMA)
'If there are things we can do to support our members, we will do what we can'
Clearly, the result of the referendum will raise many questions from the artist management community who deal with the rest of Europe on a daily basis, a common concern with all businesses in the same position. Quite what the outcome will be remains to be seen. If there are things we can do to support our members, we will do what we can – Atholl Swainston-Harrison, chief executive, International Artist Managers's Association (IAMA)