title: Molecular diversity in amino-terminal domains of human calpastatin by exon skipping.
abstract: Calpastatin, a specific inhibitor of calpain, consists of a unique N-terminal domain (domain L) and four repetitive calpain-inhibition domains (domains 1-4). Calpastatin cDNA of human was reported to have two deletions in domains L and 1, as compared with that of pig and rabbit. We isolated human calpastatin genomic DNA clones, and the sequence analysis revealed seven exons for domain L and five exons for domain 1. Those deletions in the human cDNA were retained in its genomic DNA as exons 3 and 11. By the reverse transcription polymerase chain reaction method, three calpastatin cDNAs, full-length domains L and 1, and two natural mutants with deletions in either exon 3 or in both exons 3 and 5, were cloned from human fibroblast WI-38 cell line mRNA. Domain L was found to be rich in basic amino acid residues, especially for exon 3, and its N-terminal half was highly conserved among species. The isoelectric points (pI) of domain L and domains 1-4 were calculated to be 10.27 and 4.26-4.90, respectively. Moreover, human tissues and cell lines displayed different patterns of reverse transcription polymerase chain reaction products in agarose gel electrophoresis. Therefore, alternative splicing is most likely the cause for the molecular diversity, and the multiple isoforms are implicated for specific physiological roles.
authors.affiliation: Institute for Virus Research, Faculty of Medicine, Kyoto University, Japan.
authors.forename: W J
authors.initials: WJ
authors.lastname: Lee
authors.forename: H
authors.initials: H
authors.lastname: Ma
authors.forename: E
authors.initials: E
authors.lastname: Takano
authors.forename: H Q
authors.initials: HQ
authors.lastname: Yang
authors.forename: M
authors.initials: M
authors.lastname: Hatanaka
authors.forename: M
authors.initials: M
authors.lastname: Maki
mesh_terms: D001483:Base Sequence; D002135:Calcium-Binding Proteins / Q000235:genetics*; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics; D005091:Exons; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014407:Tumor Cells, Cultured
chemical_list: D002135:Calcium-Binding Proteins; C033668:calpastatin; D004247:DNA