p27 regulates the autophagy-lysosomal pathway via the control of Ragulator and mTOR activity in amino acid deprived cells

Abstract

22 Tel: +33 (0)561558486 23 24 25 26 2 Summary 27Autophagy is a catabolic process whereby cytoplasmic components are degraded within 28 lysosomes, allowing cells to maintain energy homeostasis during nutrient depletion. Several 29 studies have shown that the CDK inhibitor p27 Kip1 promotes starvation-induced autophagy. 30 However, the underlying mechanism remains unknown. Here, we report that in amino acid 31 deprived cells, p27 controls autophagy via an mTORC1-dependent mechanism. During 32 prolonged amino acid starvation, a fraction of p27 is recruited to lysosomes where it interacts 33 with LAMTOR1, a component of the Ragulator complex required for mTORC1 lysosomal 34 localization and activation. p27 binding to LAMTOR1 prevents Ragulator assembly and function 35 and subsequent mTORC1 activation, thereby promoting autophagy. Conversely, upon amino 36 acid withdrawal, p27 -/cells exhibit elevated mTORC1 signaling, impaired lysosomal activity 37 and autophagy, and resistance to apoptosis. This is associated with sequestration of TFEB in the 38 cytoplasm, preventing the induction of lysosomal genes required for lysosomal function. 39 Silencing of LAMTOR1 or mTOR inhibition restores autophagy and induces apoptosis in p27 -/-40 cells. Together, these results reveal a direct, coordinated regulation between the cell cycle and 41 cell growth machineries. 42 43 44 45 46 47In all organisms, cell growth is coupled to cell division to allow normal development and 48 maintain homeostasis. How cells coordinate the machinery that regulates cell growth, at the heart 49 of which lies the mTOR kinase, with the machinery that controls cell division, driven by 50 cyclin/CDK complexes, has been the subject of considerable interest for several decades 1 . This 51 coordination has been mostly studied under normal metabolic conditions and except for notable 52 exceptions, such as early embryonic development, it appears that growth control drives the 53 activity of the cell cycle machinery. Here we investigated this question in conditions of 54 metabolic restriction to determine if the cell cycle machinery could in turn regulate the growth 55 control machinery. 56 p27 Kip1 (p27) was initially identified as a cyclin/CDK inhibitor 2, 3 . Due to its ability to induce 57 cell cycle arrest, p27 acts as a tumor suppressor and p27 -/mice display multiple organ 58 hyperplasia and spontaneously develop pituitary tumors 4 . Nevertheless, in contrast to classic 59 tumor suppressors such as p53 or Rb, inactivating mutations of the p27 gene are extremely rare 60 and its inactivation in cancer is rather caused by enhanced degradation, attenuated transcription 61 or translation, or mislocalization in the cytoplasm 2, 3, 5 . The latter correlates with poor prognosis 62 in a variety of cancers, suggesting a direct contribution of cytoplasmic p27 to tumor progression 2, 63 3 . In fact, knock-in mice in which p27 is largely sequestered in the nucleus due to defective 64 export to the cytoplasm (p27 S10A ) are partially resistant to tumorigene...