The Role Of Ribosomal Protein S27-like In Autophagy Of Breast Cancer Cells

Breast cancer is one of the most common malignant cancer in women.Increasing studies have shown that the deregulation of ribosomal proteins plays critical roles in the genesis and development of breast cancer.It is also well known that autophagy plays important roles in tumorigenesis.Inhibition of ribosomal biogenesis or the deletion of ribosomal proteins induces autophagy in breast cancer cells.Our previous studies showed that ribosomal protein S27-like(RPS27L),a physiological regulator of p53,suppresses tumorigenesis by maintaining genomic stability.Our immunohistochemical analysis showed that the expression of RPS27L was decreased in breast tumors,as compared to normal breast tissues.However,whether downregulation of RPS27L plays a role in breast tumorigenesis is unknown,In addition,whether RPS27L functions in a p53-independent manner to regulate autophagy is also totally unknown.To investigate the p53-independent role of RPS27L in beast tumorigenesis,we chose MB231 and SK-BR3,two breast cancer lines,harboring mutant p53.We found that siRNA-mediated silencing of RPS27L significantly induced autophagy in both breast cancer cells by immunoblotting,immunofluorescence and electron microscopy.Mechanistically,the activity of mTORC1,a major negative autophagy regulator,was significantly reduced upon RPS27L silencing,as reflected by the decreased levels of phosphorylated S6K1 and 4E-BP1,two downstream targets of mTORC1,whereas theactivity of mTORC2 was not affected,as evidenced by unchanged level of p-AKTl S473 upon RPS27L silencing.The in vitro kinase assay further confirmed that silencing of RPS27L reduced the activity of mTORCl,but had no effects on mTORC2.Furthermore,we also found that Rps271 knockout induced autophagy and decreased the activity of mTORCl in MEF cells.Finally,by western blotting and qRT-PCR analysis,we found that inactivation of RPS27L significantly upregulated the protein levels of DEPTOR,a naturally occurring inhibitor of mTOR,but slightly increased DEPTOR mRNA levels.The half-life of DEPTOR was significantly extended upon RPS27L silencing,suggesting RPS27L knockdown inhibited DEPTOR degradation,leading to its accumulation.Consistently,RPS27L silencing significantly decreased the levels ofβ-TrCP,the substrate receptor of SCF ubiquitin ligase responsible for DEPTOR degradation,by shortening its protein half-life,but has no effect on its mRNA levels.Suggesting the β-TrCP-Deptor-mTORClaxis is affected byRPS27L knockdown to induce autophagy.Biologically,blockage of RPS27L silencing induced-autophagy by chloroquine(CQ)and Bafilomycin A1(Baf-Al),two autophagy inhibitors,significantly induces apoptosis,suggesting that autophagy induced by RPS27L silencing play a protective roles in breast cancer cells.In sumary,our results suggested that RPS27L silencing triggers β-TrCP degradation via a yet-to-be identified mechanism,leading to DEPTOR accumulation to inactivate mTORC1 and induce autophagy.Autophagy induction appears to be a cellular protective mechanism for the survival of breast cancer cells.Taken together,our results suggest that the decreased expression of RPS27L may play an important role in breast tumorigenesis by triggering autophagy.Approach to inhibition autophagy in breast cancer with low RPS27L expression may,therefore,achieve a better therapeutic efficacy.Thus,our study may provide a sound rationale for personalized breast cancer treatment.