Objective:To investigate the effect of autophagy on cervical cancer cells stemness and its proliferation ability. Methods:HeLa mammospheres were isolated with the method of suspension culture. Formation of autophagosome was observed by staining with acridine orange under fluorescence microscope. Western blot was used to detect the expression of Beclin1 and LC3 B in HeLa adherent cells and mammospheres. After starvation induction with EBSS in HeLa adherent cells and mammospheres for 4 hours and 8 hours

formation of autophagosome was observed by staining with acridine orange under fluorescence microscope;Western blot was used to detect the expression of Beclin1 and LC3 B

proliferative ability of wild type HeLa cells and starvation-stimulated HeLa cells was detected using a real-time cell-based assay system. After inhibiting the autophagy with 3-methyladenine（3-MA）in HeLa adherent cells and mammospheres

Western blot was used to detect the expression of Beclin1 and LC3 B and proliferative ability of wild type HeLa cells and 3-MA-stimulated HeLa cells was detected using a real-time cell-based assay system. CRISPR/Cas9 gene editing techniques was used to down regulate the expression of Beclin1

proliferative ability of wild type HeLa cells and Beclin1-mutated HeLa cells was detected using a real-time cell-based assay system. Results:Compared with HeLa adherent cells

HeLa mammospheres had higher basic levels of autophagy. And the autophagy level was higher in HeLa mammospheres than in adherent cells under the starvation conditions. The autophagy level is significantly higher after 4 hours starvation induction both in the mammospheres and the adherent cells

which was confirmed by western blot analysis and acridine orange fluorescent staining. The proliferative ability of starvation inducted HeLa cells were weakened compared with that of wild type HeLa cells and it’s more obvious at 8 hours. The expression of Beclin1 and LC3 B are lower in the presence of 3-MA and it’s more obvious in adherent cells. The proliferative ability of HeLa cells were also weakened in the presence of 3-MA. Additionally

the proliferative ability of Beclin1-mutated HeLa cells were elevated compared with that of wild type HeLa cells. Conclusion:Autophagy is involved in maintaining the stemness of cervical cancer cells. Significantly

autophagy also can have an impact on the proliferative ability of cervical cancer cells. Gene targeting-induced disruption of autophagy key gene Beclin1 and LC3 B to alter autophagy can be a promising therapeutic strategy for curing cervical cancer.