Amino Acid Deprivation-Induced Autophagy Requires Upregulation of DIRAS3 through Reduction of E2F1 and E2F4 Transcriptional Repression.

Margie N Sutton,Zhen Lu,Jinhua Zhou,Gilbert Y Huang,Weiqun Mao,Robert C Bast,Robert Langley

doi:10.3390/cancers11050603

Abstract

Failure to cure ovarian cancer relates to the persistence of dormant, drug-resistant cancer cells following surgery and chemotherapy. “Second look” surgery can detect small, poorly vascularized nodules of persistent ovarian cancer in ~50% of patients, where >80% are undergoing autophagy and express DIRAS3. Autophagy is one mechanism by which dormant cancer cells survive in nutrient poor environments. DIRAS3 is a tumor suppressor gene downregulated in >60% of primary ovarian cancers by genetic, epigenetic, transcriptional and post-transcriptional mechanisms, that upon re-expression can induce autophagy and dormancy in a xenograft model of ovarian cancer. We examined the expression of DIRAS3 and autophagy in ovarian cancer cells following nutrient deprivation and the mechanism by which they are upregulated. We have found that DIRAS3 mediates autophagy induced by amino acid starvation, where nutrient sensing by mTOR plays a central role. Withdrawal of amino acids downregulates mTOR, decreases binding of E2F1/4 to the DIRAS3 promoter, upregulates DIRAS3 and induces autophagy. By contrast, acute amino acid deprivation did not affect epigenetic regulation of DIRAS3 or expression of miRNAs that regulate DIRAS3. Under nutrient poor conditions DIRAS3 can be transcriptionally upregulated, inducing autophagy that could sustain dormant ovarian cancer cells.

Highlights

Despite progress in surgery and chemotherapy, ovarian cancer still proves lethal in 70% of cases, leading to the death of more than 14,000 women in the United States each year [1]
When A2780 ovarian cancer cells were grown in media lacking amino acids or in serum-free Hanks buffered salt solution (HBSS)
We have found that DIRAS3 can be negatively regulated transcriptionally by binding of E2F1 and/or E2F4 to the DIRAS3 promoter, epigenetically by DNA-hypermethylation of CpG islands in promoter regions of the imprinted and non-imprinted alleles of DIRAS3, and post-transcriptionally by the expression of miRNA-221 and miRNA-222 [4,5,9]

Summary

Introduction

Despite progress in surgery and chemotherapy, ovarian cancer still proves lethal in 70% of cases, leading to the death of more than 14,000 women in the United States each year [1]. One of the most important factors contributing to poor outcomes is the persistence of dormant, drug-resistant cancer cells after primary cytoreductive surgery and combination chemotherapy. Despite normalization of CA125 and negative imaging following primary treatment, “second look” exploratory surgery of the abdominal cavity can detect small, quiescent, poorly vascularized nodules of persistent ovarian cancer on the peritoneal surface in ~50% of patients. After positive second look operations, persistent ovarian cancer may take months or years to become clinically evident, consistent with tumor dormancy. In more than 80% of positive second looks, persistent ovarian cancer cells express DIRAS3 and are undergoing autophagy [2].

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Conclusion