P.164Expression of alternative nebulin isoforms containing super repeat S21a or S21b in skeletal muscle

Abstract

Nebulin is a giant sarcomeric protein, binding to actin along the length of the thin filament. Mutations in NEB are the most common cause of autosomal recessive nemaline myopathy (NM). NEB has 183 exons, and undergoes extensive alternative splicing, giving rise to numerous different protein isoforms. The protein structure is modular, with 32-35 amino-acid actin-binding simple repeats forming 26 different super repeats spanning most of the protein. Two alternative isoforms of super repeat S21, S21a and S21b, are encoded by two mutually exclusive exons, exon 143 and exon 144, respectively. S21a and S21b differ in both charge and hydrophobicity. The amino-acid sequence encoded by exon 143 shows complete homology between mouse, rat and human, indicating a central, conserved role for this region of the nebulin protein. To study the expression and function of these developmentally and clinically intriguing alternatively spliced exons, we have developed specific monoclonal antibodies, NEB-143 and NEB-144, against the isoforms containing S21a or S21b, respectively. We have previously shown that nebulin containing exon 144 is the first isoform expressed in myogenesis in vitro, while regulated expression of nebulin containing exon 143 occurs at later stages of muscle development. In this study, we investigated the expression of the S21a (exon 143) nebulin isoform further, in several adult muscles and myofibre types. Our results revealed an interesting pattern of alternating correlation between the expression of S21a and different types of MyHC myofibres, both slow and fast, with preferential expression in fast, MyHC IIX fibres.