A Novel Mutation of the Spectrin Gene
A Novel Mutation of the Spectrin Gene
The purpose of this study is to find a gene or its mutation (an altered gene) that puts
 individuals at risk for developing HE or HPP.
Mutations of spectrin (Sp) involving the Sp heterodimer self-association site (the I domain
 of Sp) represent the most common group of membrane skeletal defects in hereditary
 elliptocytosis (HE) and a closely related disorder, hereditary pyropoikilocytosis (HPP). HPP
 is characterized by extreme anisocytic microcytosis, a severe spectrin dimer self-association
 defect and spectrin deficiency. The conventional explanation for the different phenotypes is
 that HPP subjects are compound heterozygotes for an sp defect that interferes with sp
 tetramer assembly and a second defect which results in decreased synthesis of functional sp.
 In contrast, HE subjects have normal spectrin content and a less severe sp self-association
 defect. The clinical expression of HE/HPP is influenced by the inheritance of modifying
 factors such as the Sp hypomorphic mutation, LELY. LELY is a low expression allele of the Sp
 gene characterized by a C>G mutation in codon 1857 of exon 40 and a C>T -12 mutation in
 intron 45 that is responsible for partial skipping of exon 46, which is essential for the
 functional assembly of /b sp dimers.

 CASE REPORT:

 Here we describe four members from a family of northern European descent in which members had
 different erythrocyte morphology ranging from atypical HPP to HE to normal and a novel Sp
 mutation. Quantitation of RBC membrane proteins of the propositus with atypical
 non-microcytic HPP revealed 48% spectrin dimers (control 10%) due to a marked increase in the
 74kD I Sp peptide. There was only a slight decrease in the spectrin/band 3 ratio, which
 correlated with the normocytic morphology. There was also an abnormal Sp peptide at 41kD
 suggesting presence of LELY. Sequencing of his Sp gene revealed heterozygosity for a novel
 mutation in exon 2, codon 34: CGG->CCG (Arg>Pro) and heterozygosity for LELY. These mutations
 were also present in his brother and daughter who have HE, while another son, who had
 Arg34Pro, but not LELY has repeatedly confirmed normal morphology.

 We plan to screen 11 other family members who are suspected to have HPP or HE. Blood samples
 will be obtained and the following will be performed:

 1. Quantitation of erythrocyte (RBC) membrane proteins

 2. DNA extraction from the WBC and screening for spectrin mutation with restriction enzymes
 (if needed we will sequence spectrin gene)

 3. Quantitative Real Time PCR testing will be done on RNA to assess the spectrin gene
 expression

 4. History, Clinical examination, CBC, chemistry and peripheral blood smear

 Quantitation of RBC membrane proteins will be done by looking for the abnormal increase in
 the amount of spectrin dimers in patients with HPP (control 10%). We will also look at the
 DNA for any known or yet unreported mutations in the spectrin gene and we will correlate the
 expression of spectrin gene wrt level of spectrin protein in the RNA.

 No statistical analysis will be done. This is an observation study designed to correlate
 clinical features with the above mentioned tests results.
Inclusion Criteria:

 -  > 7 years of age

 -  Consenting family members

 Exclusion Criteria:

 -  Anyone not meeting the criteria above