Cdna Cloning and Expression Analysis of Chaperonin Containing T-Complex Polypeptide 1-Alpha in the Pearl Oyster Pinctada maxima Exposed to Cold Stress

Abstract

ABSTRACT The complete cDNA sequence of chaperonin-containing T-complex polypeptide 1-alpha (CCT-alpha) gene of the pearl oyster Pinctada maxima was determined using rapid amplification of cDNA ends technique and characterized with bioinformatic analysis. Its expression profile in different organs and under cold stress was characterized using real-time quantitative polymerase chain reaction. Results revealed that the complete cDNA sequence of P. maxima CCT-alpha has 1,803 base pairs (bp), including a 1,608-bp open reading framework that codes 535 amino acids. The protein has three conservative functional domains: equatorial, apical, and intermediate domains. The equatorial domain is 20 amino acids shorter than that of other species, and contains the three signature sequences of CCT family protein. The CCT-alpha of P. maxima has nine conservative ATP-binding sites and six protein-binding sites, and has high homology with Aplysia californica (80.7%), Delia antiqua (77.4%), Danio rerio (77.1%), Xenopus laevis (76.7%), Gallus gallus (76.4%), and Homo sapiens (75.4%). The CCT-alpha shows tissue-specific expression profile in P. maxima, with expression levels higher in gill, mantle, and adductor muscle than in hepatopancreas and heart. Cold stress markedly increased mRNA contents of CCT-alpha in the organs of P. maxima, but different organs showed distinct cold stress response pattern. Chaperonin-containing T-complex polypeptide 1-alpha transcription in adductor muscle, hepatopancreas, and heart increased at 2 h, and reached peak values at 6 h, maintained high levels at 12 h, and significantly decreased at 24 h. The product in gill and mantle decreased at 2 h, began to increase at 6 h, and showed a large increase at 12 h, but reduced to below the initial level at 24 h.