Gene cloning, sequence analysis and pro-apoptotic functional study on large yellow croaker Bmf2

Bcl-2 modifying factor (Bmf), a member of Bcl-2 family bearing only BH3 domain, can initiate apoptosis in eukaryotic cells when apoptotic stimulation signals exist. It plays an important role in embryonic development, organogenesis and tumor suppression. Currently, Bmf from lower vertebrates has only been reported in zebrafish. Sequence alignment, gene synteny and phylogenetic analysis showed that there are two different Bmf genes, Bmf1 and Bmf2, in teleost. In this study, we cloned an open reading frame sequence of large yellow croaker Bmf2 (LcBmf2), which is 561 bp long and encoded 187 amino acids. Although the sequence identities between LcBmf2 and human or mouse Bmfs were low, LcBmf2 contains the DLC2 binding motif and conserved BH3 domain, which are highly important to Bmf function, suggesting it might exhibit similar functions to its mammalian homologue. The transient overexpression of LcBmf2 in HEK-293T cells induced cell detachment, morphology change as well as the intracellular enzyme activity increase of Caspase 3 and Caspase 8, indicating LcBmf2 prompted apoptosis in HEK-293 T cells. This is the first report on the pro-apoptotic function of a teleost Bmf at cellular and molecular level, which lays the foundation for further research on the role and molecular mechanism of Bmf in teleost cell apoptosis.