Twelve actin-encoding cDNAs from the American lobster, Homarus americanus: Cloning and tissue expression of eight skeletal muscle, one heart, and three cytoplasmic isoforms

Abstract

Lobster muscles express a diverse array of myofibrillar protein isoforms. Three fiber types (fast, slow-twitch or S1, and slow-tonic or S2) differ qualitatively and quantitatively in myosin heavy and light chains, troponin- T, -I, and -C, paramyosin, and tropomyosin variants. However, little is known about the diversity of actin isoforms present in crustacean tissues. In this report we characterized cDNAs that encode twelve actin isoforms in the american lobster, Homarus americanus: eight from skeletal muscle (Ha-ActinSK1-8), one from heart (Ha-ActinHT1), and three cytoplasmic type actins from hepatopancreas (Ha-ActinCT1-3). All twelve cDNAs were products of distinct genes, as indicated by differences in the 3′-untranslated regions (UTRs). The open reading frames specified polypeptides 376 or 377 amino acids in length. Although key amino residues are conserved in the lobster actins, variations in nearby sequences may affect actin polymerization and/or interactions with other myofibrillar proteins. Quantitative reverse transcriptionpolymerase chain reaction showed muscle fiber type- and tissue-specific expression patterns. Ha-Actin-HT1 was expressed exclusively in heart (87% of the total;12% of the total was Ha-ActinCT1). Ha-ActinCT1 was expressed in all tissues, while CT2 and CT3 were expressed only in hepatopancreas, with Ha-ActinCT2 as the major isoform (93% of the total). Ha-ActinSK1 and SK2 were the major isoforms (88% and 12% of the total, respectively) in the S1 fibers of crusher claw closer muscle. Fast fibers in the cutter claw closer and deep abdominal muscles differed in SK isoforms. Ha-ActinSK3, SK4, and SK5 were the major isoforms in cutter claw closer muscle (12%, 48%, and 37% of the total, respectively). Ha-ActinSK5 and SK8 were the major isoforms in deep abdominal flexor (31% and 65% of the total, respectively) and extensor (46% and 53% of the total, respectively) muscles, with SK6 and SK7 expressed at low levels. These data indicate that fast fib