Correct NH2-terminal processing of cardiac muscle alpha-isoactin (class II) in a nonmuscle mouse cell.
Both mammalian nonmuscle and muscle actins possess an AcAsp(Glu)NH2 terminus. The nonmuscle actin genes code for a polypeptide with a Met-Asp NH2 terminus (class I) whereas the muscle actin genes code for a polypeptide with a Met-Cys-Asp NH2 terminus (class II). Two amino acids must be removed for mature muscle actin synthesis, whereas only the Met must be removed for nonmuscle actin synthesis. We wished to know whether a nonmuscle cell which normally does not synthesize a class I actin can correctly process a muscle actin with its extra NH2-terminal amino acid in vivo. To answer this question we have used L/LK165 cells, a mouse L-cell transfected with a human cardiac muscle actin gene. When these cells were labeled overnight with [35S]Cys, an actin with an NH2-terminal tryptic peptide corresponding to that of mature cardiac muscle actin was detected. When the cells were pulse-labeled for 20 min, a new actin intermediate containing an AcCys-Asp amino terminus was observed which then disappeared with time. Furthermore, the muscle actin was processed as fast if not faster than the nonmuscle actin in these cells. This actin intermediate was also seen in chick myotube cultures. Our results show that the ability to correctly process muscle specific actins is not tissue specific. Furthermore, these results confirm a processing pathway for class II actins proposed by us earlier on the basis of experiments with a cell-free translation system.
D000199:Actins / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000032:analysis; D002460:Cell Line; D003545:Cysteine / Q000032:analysis; D005796:Genes; D006801:Humans; D007739:L Cells / Q000378:metabolism; D008715:Methionine / Q000032:analysis; D051379:Mice; D009206:Myocardium / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D013820:Thermolysin; D014357:Trypsin
D000199:Actins; D010446:Peptide Fragments; D001224:Aspartic Acid; D008715:Methionine; D014357:Trypsin; D013820:Thermolysin; D003545:Cysteine