Two isoforms of smooth muscle myosin regulatory light chain in chicken gizzard

We isolated a cDNA clone for a new isoform of chicken smooth muscle myosin regulatory light chain (MRLC) from a cDNA library of embryonic chicken gizzard. The deduced amino acid sequence was different in 10 amino acid residues from the previously reported polypeptide sequences of chicken gizzard MRLC. The in vitro transcription/translation product from the cDNA comigrated with a minor isoform of chicken gizzard MRLC (L20-B) in a two-dimensional gel electrophoresis. This isoform was detected only in the embryonic gizzard and was slightly more acidic than the predominant isoform (L20-A). The partial polypeptide sequence of L20-A was confirmed to be identical to the previously reported MRLC sequence. Nevertheless, Northern blot analysis showed that L20-B-related mRNAs were present in both the embryonic and adult gizzard. Non-denaturing pyrophosphate polyacrylamide gel electrophoresis showed that the in vitro transcription/translation product could be associated with native myosin when mixed and coprecipitated in a low-ionic-strength buffer with adult chicken gizzard myosin. Moreover, the coprecipitated translation product was phosphorylated in vitro by chicken gizzard myosin light chain kinase apparently more rapidly than L20-A on the native myosin heavy chain. From these findings, we concluded that at least two isoforms of smooth muscle MRLC exist in chicken gizzard and that their expression may be regulated translationally depending on the developmental stage.