ULK1·ATG13·FIP200 Complex Mediates mTOR Signaling and Is Essential for Autophagy

Abstract

Autophagy is a degradative process that recycles long-lived and faulty cellular components. It is linked to many diseases and is required for normal development. ULK1, a mammalian serine/threonine protein kinase, plays a key role in the initial stages of autophagy, though the exact molecular mechanism is unknown. Here we report identification of a novel protein complex containing ULK1 and two additional protein factors, FIP200 and ATG13, all of which are essential for starvationinduced autophagy. Both FIP200 and ATG13 are critical for correct localization of ULK1 to the pre-autophagosome and stability of ULK1 protein. Additionally, we demonstrate by using both cellular experiments and a de novo in vitro reconstituted reaction that FIP200 and ATG13 can enhance ULK1 kinase activity individually but both are required for maximal stimulation. Further, we show that ATG13 and ULK1 are phosphorylated by the mTOR pathway in a nutrient starvation-regulated manner, indicating that the ULK1⅐ATG13⅐FIP200 complex acts as a node for integrating incoming autophagy signals into autophagosome biogenesis.Macroautophagy (herein referred to as autophagy) is a catabolic process whereby long-lived proteins and damaged organelles are shuttled to lysosomes for degradation. This process is conserved in all eukaryotes. Under normal growth conditions a housekeeping level of autophagy exists. Under stress, such as nutrient starvation, autophagy is strongly induced resulting in the engulfment of cytosolic components and organelles in specialized double-membrane structures termed autophagosomes. Following fusion of the outer autophagosomal membrane with lysosomes, the inner membrane and its cytoplasmic cargo are degraded and recycled (1-3). Recent work has implicated autophagy in many disease pathologies, including cancer, neurodegeneration, as well as in eliminating intracellular pathogens (4 -8).The morphology of autophagy was first described in mammalian cells over 50 years ago (9). However, it is only recently through yeast genetic screens, that multiple autophagy-related (ATG) genes have been identified (10 -12). The yeast ATG proteins have been classified into four major groups: the Atg1 protein kinase complex, the Vps34 phosphatidylinositol 3-phosphate kinase complex, the Atg8/Atg12 conjugation systems, and the Atg9 recycling complex (13). Even though many ATG genes are now known, most of which have functional homologs in mammalian cells (14, 15), the molecular mechanism by which they sense the initial triggers and subsequently dictate autophagy-specific intracellular membrane events is far from understood.In yeast, one of the earliest autophagy-specific events is believed to involve the Atg1 protein kinase complex. Atg1 is a serine/threonine protein kinase and a key autophagy-regulator (16). Atg1 is complexed to at least two other proteins during autophagy, Atg13 and Atg17, both of which are required for normal Atg1 function and autophagosome generation (17-19). Classical signaling pathways such as the cAMP-dependent kinase (PKA) pat...

Hexokinase II integrates energy metabolism and cellular protection: Akting on mitochondria and TORCing to autophagy

“…Consequently, ULK1 is activated and initiates membrane nucleation, which is followed by sequestration and degradation. [155][156][157][158][159][160] Although it has been widely accepted that autophagy is induced in response to metabolic suppression, the molecular links between metabolic and autophagic pathways have not been fully elucidated. Recently, we made the surprising discovery that HK-II facilitates autophagy in response to glucose deprivation to protect cells 117 (Figure 4).…”

mTOR-Dependent Cell Survival Mechanisms

“…Recently, several new substrates for mTORC1 have been identified and characterized. For example, mTORC1 also controls growth by negatively regulating autophagy through the direct phosphorylation of Unc-51-like kinase (ULK1; discussed below) (Chan 2009;Ganley et al 2009;Hosokawa et al 2009;Jung et al 2009). In addition, mTORC1 directly phosphorylates the phosphatidate phosphatase Lipin1 (which regulates lipid metabolism through SREBP1) and the growth factor receptor-bound protein 10 (Grb10; part of a negative feedback loop targeting insulin receptor signaling) (Hsu et al 2011;Peterson et al 2011;Yu et al 2011).…”

Fission yeast Ryh1 GTPase activates TOR Complex 2 in response to glucose

“…6,[23][24][25][26][27][28][29][30][31] Fission yeast TORC1 is activated in the presence of nitrogen and carbon sources, and phosphorylates the major S6 kinase Psk1. 22,[32][33][34] In this organism, TORC2 is also implicated in cellular nutritional responses, including the G1 arrest upon nitrogen starvation and the mitotic control in response to glucose.…”