Structure of mouse calpastatin isoforms: Implications of species-common and species-specific alternative splicing

Mouse calpastatin cDNAs were cloned by the method of RT-PCR using RNA isola ted from myoblast C2C12 cells. Nucleotide sequencing of the isolated clones revealed an in-frame ATG codon upstream of the previously assigned transla tion initiation methionine. Except for the N-terminal segment, the new tran slatable region (domain XL) was similar to the sequence of bovine calpastat in in which domain XL was first identified. Among the isolated mouse calpas tatin cDNA clones, three isoforms (mCS-a, mCS-b, and mCS-c) were identified . In domain L, mCS-b had a deletion of the region corresponding to exon 3 o f the human calpastatin gene. RT-PCR analyses of various mouse tissues reve aled that mCS-b was the major form and that the content of mCS-a, nondelete d form, was 5-10% in tissues including skeletal muscle, liver, brain, etc. and about 30% in the myoblast C2C12 cells. Unlike human and rat cDNAs, no o ther deletions were detected in mouse calpastatin domain L. Isolation of th e cDNA clone of mCS-c, which lacked regions corresponding to exons 3 and 12 , was obtained by chance because its expression level was under the detecta ble level in the mouse tissues and even in C2C12 cells. (C) 1999 Academic P ress.