BETA-SPECTRIN CAMPINAS - A NOVEL SHORTENED BETA-CHAIN VARIANT ASSOCIATED WITH SKIPPING OF EXON-30 AND HEREDITARY ELLIPTOCYTOSIS

beta-Spectrin Campinas is a novel spectrin variant associated with a s hortened beta-chain in a kindred with hereditary elliptocytosis (HE). The propositus and her mother exhibited increased amounts of spectrin dimers and an increase in the alpha I 74 kD fragment from the alpha-ch ain after partial tryptic digestion of spectrin. The shortened beta-ch ain appeared as an additional band of approximately 200kD on SDS-PAGE. In order to delineate the molecular defect of this abnormality at the gene level, reticulocyte mRNA was transcribed into cDNA and the last four exons of the beta-spectrin gene were amplified. Agarose gel of th e amplification product of the propositus revealed the expected band o f 487 bp as well as a shortened band of approximately 300 bp (size det ermined on gel). This shortened cDNA amplification product was cloned and nucleotide sequencing revealed the absence of the entire exon 30. In order to determine the underlying mutation responsible for this abn ormal splicing, a genomic DNA fragment containing exons 30 and 31 was amplified and nucleotide sequencing revealed a G --> A substitution at the 5' donor splice site consensus sequence of intron 30 (nt+1 IVS30) . The skip splicing observed in this study results in a frameshift, cr eating a new stop codon and causing a deletion of 129 aminoacids at th e very COOH-terminus of the protein, thus impairing spectrin dimers se lf-association. We classified this HE as spherocytic HE because the pr opositus presented a few spherocytes in addition to many elliptocytes in the blood smear, whereas her mother, who was splenectomized, showed many schizocytes, poikilocytes and spherocytes.