Objective: To investigate the role of Beclin 1 in HeLa cells and to obtain further insight into the relationship between autophagy and apoptosis.
Methods: Beclin 1 silencing was achieved using RNA interference. The expression of gene was measured using quantitative real time RT-PCR and Western blotting. The percentage of apoptotic cells and cell cycle analysis and cell proliferation were assessed by flow cytometry and MTT assay. The ultrastructural analysis was under the electron microscope.
Results: In pSUPER-Bec transfectants (Beclin 1 gene partially silenced) the expression of mRNA and protein of Beclin 1 were significantly suppressed in comparison to pSUPER-non (scramble RNA control) or untreated cells in HeLa cells. The growth of transfected cells was promoted, and less apoptosis cells were identified in pSUPER-Bec transfectants compared with pSUPER-non transfectants. Meanwhile pcDNA3.1-Bec transfectants (Beclin 1 gene overexpressed) showed reduction of cell proliferation but augmentation of cell programmed death compared with vector vehicle. The autophagy-promoting activity of beclin 1 in HeLa cells is associated with inhibition of HeLa cellular proliferation, in vivo tumorigenesis in nude mice. The expression pattern of caspase-9 was extraordinarily similar to that of Beclin 1in siRNA against Beclin 1 transfectants and constructive expression of Beclin 1transfectants.
Conclusion: siRNA against Beclin 1 transfectants promoted the cell proliferation but overexpression of Beclin 1 promoted the autophagy cell death, and in the process of autophagy triggered by Beclin 1 expression followed accordingly the regulation of the expression of caspase-9. We conjecture that the autophagy gene Beclin 1 may be the critical molecular switch that plays an important role in fine tuning the autophagy and apoptosis through caspase-9, and defection of autophagy or apoptosis may be an important mechanism in tumorigenesis.