ChBax/Bak as key regulators of the mitochondrial apoptotic pathway: Cloned and characterized in Crassostrea hongkongensis

• The cDNA sequences of ChBax/Bak were first cloned from Hong Kong oyster Crassostrea hongkongensis.
• Expression levels of ChBax/Bak changed during the embryonic development and under the challenge of microorganisms.
• Over-expression of ChBax/Bak in HEK293 T cells active the transcriptional activities of p53/p21-Luc reporter gene.

Apoptosis has been primarily investigated in mammals, and little is known about apoptosis in mollusks. The proteins Bax and Bak play critical roles in the mitochondrial apoptosis pathway and in determining cell fate. In this study, ChBax and ChBak, homologs of the well-known Bax and Bak proteins, were identified from the oyster Crassostrea hongkongensis. The ChBax/Bak proteins consist of 207/232 amino acids with the typical domains found in BCL-2 family members. ChBax and ChBak mRNA expression were detected in all 8 of the selected oyster tissues and at the different stages of development. Fluorescence microscopy revealed that the full-length proteins of ChBax/Bak were located in the cytoplasm and mitochondrial outer membrane, of HEK293T cells, respectively. Furthermore, both of the genes' expression levels were found to increase in the hemocytes of oysters challenged with pathogens. The over-expression of ChBax or ChBak activates the p53-Luc reporter gene in HEK293T cells in a dose-dependent manner. These results indicate that ChBax and ChBak may play important roles in the mitochondrial apoptotic pathway in oysters.