Proteotoxic and metabolic stress crosstalk in colon cancer: role of Sestrin 2

Proteome homeostasis is fundamental for cell health. Organisms respond to proteotoxic stress (PS) by activating the transcription factor HSF1, which regulates a PS-driven transcriptional response inducing the expression of cytoprotective heat shock genes. The natural cyclopentenone 15-deoxyΔ 12,14 -prostaglandin J2 (15d-PGJ2), which possesses a unique spectrum of biological activities, is known to be a potent inducer of HSF1 activation. Recently, the study of the biological activity of this prostanoid in our laboratory has led to the discovery of two new human heat shock genes, AIRAP and NKRF, whose expression is regulated by HSF1. In addition to these genes, a third gene, Sestrin 2 (SESN2), was found to be highly expressed in a microarray analysis of gene expression profile of HeLa cells treated with 15d-PGJ2. SESN2, a highly conserved stress-responsive protein, plays an important role in cell metabolism through the inhibition of the mechanistic target of rapamycin complex 1 (mTORC1), an important positive regulator of cell growth and proliferation. Based on the initial microarray analysis observation, this study was undertaken to: 1. confirm the ability of 15d-PGJ2 to induce SESN2 expression in cancer cells; 2. investigate the molecular mechanism responsible for induction of SESN2 expression by 15d-PGJ2; 3. analyze the role of HSF1 in SESN2 gene expression; 4. investigate the possible role of SESN2 and the mTOR pathway in the anticancer activity of 15d-PGJ2. The results obtained demonstrate that 15d-PGJ2 is a potent inducer of SESN2 expression in different types of cancers, including lung, breast and colon carcinoma, triggering de novo SESN2 transcription. The regulation of SESN2 gene expression is very complex and it is still not well understood. In silico analysis of the human SESN2 promoter confirmed the presence of binding sites for several transcription factors previously shown to regulate SESN2 gene expression, including p53, Nrf2 and ATF4; however, none of these factors was found to regulate 15d-PGJ2-induced SESN2 transcription. Instead this study identifies the Akt pathway as a critical regulator of 15dPGJ2-induced SESN2 expression in cancer cells. Interestingly, the in silico SESN2 promoter analysis led to the identification of a heat shock element (HSE3) able to bind HSF1 in the SESN2 promoter. Surprisingly, the investigation on HSF1 function in 15d-PGJ2-treated cells identified HSF1 as an important negative regulator of SESN2. In the second part of my work, the study of the function of 15d-PGJ2-induced SESN2 expression led to the demonstration of a potent antitumor activity of 15d-PGJ2 in colon cancer. More importantly, we identified the natural prostanoid 15d-PGJ2 as a novel inhibitor of the mTORC1 pathway, an effect dependent on SESN2 expression. The results highlight the important role of the Akt signaling pathway in SESN2 expression, and demonstrate, for the first time, a regulatory role of HSF1 on the human SESN2 gene, opening new perspectives in the understanding of the role of the proteostasis guardian HSF1 in SESN2-mediated cell metabolism regulation in cancer cells.