Calponin Isoform Expression in the Japanese Pearl Oyster, <i>Pinctada fucata</i>

Abstract

Calponin is a basic actin-binding protein found widely in invertebrate tissues including catch muscle and therefore may participate in catch contraction. There is limited information about molluscan calponin and molecular characterization to reveal its function in the regulatory system. We previously identified and partially sequenced three calponin isoforms of the Japanese pearl oyster, Pinctada fucata (Pifuc-CP-1, Pifuc-CP-2 and Pifuc-CP-3). In this study, the full-length nucleotide sequences of the three isoforms were determined. The primary structures revealed that Pifuc-CP-1 consists of 324 amino acids (aa) with a molecular mass (Mw) of 34.7 kDa and an isoelectric point (pI) of 9.40. Pifuc-CP-2 is 303 aa in length with a Mw of 33.3 kDa and a pI of 9.30, and Pifuc-CP-3 is 398 aa in length with a Mw of 43.8 kDa and a pI of 8.55. Domain architecture prediction showed that the three isoforms have a single calponin homology (CH) domain and multiple calponin (CN) domains. Pifuc-CP-1, Pifuc-CP-2 and Pifuc-CP-3 possess four, three and five CN domains, respectively. Tissue distribution analysis indicated the presence of additional calponin isoforms and these isoforms are distributed widely in muscle and non-muscle tissues. Results of cDNA cloning revealed further four calponin isoforms: Pifuc-CP-4 (402 aa, 42.8 kDa, pI = 9.10), Pifuc-CP-5 (285 aa, 30.7 kDa, pI = 9.45), Pifuc-CP-6 (286 aa, 31.1 kDa, pI = 9.60) and Pifuc-CP-7 (302 aa, 33.3 kDa, pI = 9.10). The domain architecture of these four isoforms also consists of a single CH domain and multiple CN domains. Pifuc-CP-4 possesses six CN domains, whereas Pifuc-CP-5, Pifuc-CP-6 and Pifuc-CP-7 contain three CN domains. Sequence alignment of P. fucata calponin isoforms showed that Pifuc-CP-1, Pifuc-CP-2, Pifuc-CP-3 and Pifuc-CP-4 have identical CH domain sequences, whereas Pifuc-CP-5, Pifuc-CP-6 and Pifuc-CP-7 have identical CH domain sequences. The CN repeats were not well conserved. These findings suggest that P. fucata calponin isoforms function differently in each tissue.

Troponin T from the Japanese Pearl Oyster <i>Pinctada fucata</i>: Molecular Cloning, Tissue Distribution, Gene Structure, and Interaction Analysis with Tropomyosin

“…In summary, our group has now described the molecular characteristics of all of the key proteins thought to be involved in the thin filament-linked regulatory system of P. fucata: troponin C [14], troponin I [15], troponin T (this study), tropomyosin-1 and -2 [24], and calponin-1, -2, -3, -4, -5, -6 and -7 [31]. Calponin, a thin filament-associated protein, is considered to be involved in vertebrate smooth muscle contraction.…”

A Preparative Method for the Isolation of Calponin from Molluscan Catch Muscle

“…The molecular mechanism underlying latch contraction remains unclear [ 10 ]; however, it has been suggested that calponin is involved in the tethering of thick- and thin-filaments, like molluscan twitching [ 11 ]. The latch state of vertebrate smooth muscles is similar to the catch state of molluscan smooth muscles [ 11 ], which also contain calponin-like proteins [ 12 , 13 ].…”

“…Calponin can influence the ATPase cycle by changing the actin conformation. Calponin may be involved in the regulation of catch muscle contraction in molluscs [ 10 ].…”

“…Studies on the molecular genetics of catch muscle calponin have revealed the full length complementary DNA sequence from Mizuhopecten yessoensis [ 17 ], Crenomytilus grayanus [ 18 ], and the pearl oyster Pinctada fucata [ 10 ]. Studies have investigated the tissue expression of these proteins, including the distribution and number of isoforms [ 10 , 19 ]. Isoforms of calponin at 34 and 40 kDa have been identified in the catch muscle of the mussel C. grayanus [ 13 ].…”