Abstract:
The change of intracellular calcium (Ca
2+) concentration is an important factor in oocyte maturation. As a member of the P-type ATPase family and a crucial regulator of intracellular calcium transport, sarco/endoplasmic reticulum calcium adenosine triphosphatase (SERCA) is a key enzyme. To investigate the function of SERCA in the development of the oocytes of
Sipunculus nudus, we obtained the full length of
S. nudus SERCA (
Sn-SERCA) cDNA by using RACE technique, and determined the
Sn-SERCA relative expression level in different oocyte developmental periods by using real-time fluorescence quantitative PCR (qRT-PCR). The results indicate that the overall length of the
Sn-SERCA was 3 840 bp, the 5'UTR was 196 bp, the 3'UTR was 581 bp, and the open reading frame was 3 060 bp, encoding 1 020 amino acids. Sn-SERCA had TGES and DKTGT which were the P-type ATPase family's two conserved motifs and were necessary for catalytic reactions. According to multiple sequence alignment, motif analysis, and predictions of tertiary structures, the SERCA homologous proteins exhibited great conservatism. The phylogenetic tree analysis shows that the Sn-SERCA formed a broad branch with homologous protein sequences from invertebrates such as
Lumbricus rubellus and
Lingula anatine. The result of qRT-PCR demonstrates that the
Sn-SERCA was significantly expressed in the late yolk vigorous synthesis period and had the maximum value during coelomic fluid development. When the oocytes moved to the nephridioduct, the
Sn-SERCA expression was considerably higher than that at other time (
P<0.05). The variable expression of
Sn-SERCA in different periods of oocyte development suggests that Sn-SERCA is crucial in the development and germinal vesicle breakdown in
S. nudus oocytes.