ABSTRACT The AMP-activated serine/threonine protein kinase (AMPK) is a sensor of cellular energy ... more ABSTRACT The AMP-activated serine/threonine protein kinase (AMPK) is a sensor of cellular energy status found in all eukaryotes—that is, activated under conditions of low-intracellular adenosine triphosphate (ATP) following cellular stress. One of the major downstream signaling pathways regulated by AMPK is the mammalian target-of-rapamycin (mTOR pathway). Like AMPK, the mTOR serine/threonine kinase plays key roles in growth control and cell proliferation and also in metabolism. Across eukaryotes mTOR orthologs are found in two biochemically distinct complexes and only one of those complexes (mTORC1 in mammals) is acutely sensitive to rapamycin and regulated by nutrients and AMPK. AMPK directly phosphorylates at least two proteins to induce rapid inhibition of mTORC1 activity, the tuberous sclerosis protein 2 (TSC2) tumor suppressor and the mTOR binding partner raptor. The chapter explores the molecular connections between AMPK and mTOR signaling pathways and examines the physiological processes in which AMPK regulation of mTOR is critical for growth or metabolic control. The AMP-activated protein kinase (AMPK) is a heterotrimeric kinase complex composed of a catalytic (α) subunit and two regulatory (β and γ) subunits.
Adenosine monophosphate-activated protein kinase (AMPK) is a conserved sensor of intracellular en... more Adenosine monophosphate-activated protein kinase (AMPK) is a conserved sensor of intracellular energy activated in response to low nutrient availability and environmental stress. In a screen for conserved substrates of AMPK, we identified ULK1 and ULK2, mammalian orthologs of the yeast protein kinase Atg1, which is required for autophagy. Genetic analysis of AMPK or ULK1 in mammalian liver and Caenorhabditis elegans revealed a requirement for these kinases in autophagy. In mammals, loss of AMPK or ULK1 resulted in aberrant accumulation of the autophagy adaptor p62 and defective mitophagy. Reconstitution of ULK1-deficient cells with a mutant ULK1 that cannot be phosphorylated by AMPK revealed that such phosphorylation is required for mitochondrial homeostasis and cell survival during starvation. These findings uncover a conserved biochemical mechanism coupling nutrient status with autophagy and cell survival.
Background: The appropriate control of mitotic entry and exit is reliant on a series of interlock... more Background: The appropriate control of mitotic entry and exit is reliant on a series of interlocking signaling events that coordinately drive the biological processes required for accurate cell division. Overlaid onto these signals that promote orchestrated cell division are checkpoints that ensure appropriate mitotic spindle formation, a lack of DNA damage, kinetochore attachment, and that each daughter cell has the appropriate complement of DNA. We recently discovered that AMP-activated protein kinase (AMPK) modulates the G2/M phase of cell cycle progression in part through its suppression of mammalian target of rapamycin (mTOR) signaling. AMPK directly phosphorylates the critical mTOR binding partner raptor inhibiting mTORC1 (mTOR-raptor rapamycin sensitive mTOR kinase complex 1). As mTOR has been previously tied to mitotic control, we examined further how raptor may contribute to this process.
AMPK is a highly conserved sensor of cellular energy status that is activated under conditions of... more AMPK is a highly conserved sensor of cellular energy status that is activated under conditions of low intracellular ATP. AMPK responds to energy stress by suppressing cell growth and biosynthetic processes, in part through its inhibition of the rapamycin-sensitive mTOR (mTORC1) pathway. AMPK phosphorylation of the TSC2 tumor suppressor contributes to suppression of mTORC1, however TSC2-deficient cells remain responsive to energy stress. Using a proteomic and bioinformatics approach, we sought to identify additional substrates of AMPK that mediate its effects on growth control. We report here that AMPK directly phosphorylates the mTOR binding partner raptor on two well-conserved serine residues, and this phosphorylation induces 14-3-3 binding to raptor. The phosphorylation of raptor by AMPK is required for the inhibition of mTORC1 and cell cycle arrest induced by energy stress. These findings uncover a novel conserved effector of AMPK that mediates its role as a metabolic checkpoint coordinating cell growth with energy status.
Resveratrol (3,5,49-trihydroxystilbene) extends the lifespan of diverse species including Sacchar... more Resveratrol (3,5,49-trihydroxystilbene) extends the lifespan of diverse species including Saccharomyces cerevisiae, Caenorhabditis elegans and Drosophila melanogaster. In these organisms, lifespan extension is dependent on Sir2, a conserved deacetylase proposed to underlie the beneficial effects of caloric restriction. Here we show that resveratrol shifts the physiology of middle-aged mice on a high-calorie diet towards that of mice on a standard diet and significantly increases their survival. Resveratrol produces changes associated with longer lifespan, including increased insulin sensitivity, reduced insulin-like growth factor-1 (IGF-I) levels, increased AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-c coactivator 1a (PGC-1a) activity, increased mitochondrial number, and improved motor function. Parametric analysis of gene set enrichment revealed that resveratrol opposed the effects of the high-calorie diet in 144 out of 153 significantly altered pathways. These data show that improving general health in mammals using small molecules is an attainable goal, and point to new approaches for treating obesity-related disorders and diseases of ageing.
ABSTRACT The AMP-activated serine/threonine protein kinase (AMPK) is a sensor of cellular energy ... more ABSTRACT The AMP-activated serine/threonine protein kinase (AMPK) is a sensor of cellular energy status found in all eukaryotes—that is, activated under conditions of low-intracellular adenosine triphosphate (ATP) following cellular stress. One of the major downstream signaling pathways regulated by AMPK is the mammalian target-of-rapamycin (mTOR pathway). Like AMPK, the mTOR serine/threonine kinase plays key roles in growth control and cell proliferation and also in metabolism. Across eukaryotes mTOR orthologs are found in two biochemically distinct complexes and only one of those complexes (mTORC1 in mammals) is acutely sensitive to rapamycin and regulated by nutrients and AMPK. AMPK directly phosphorylates at least two proteins to induce rapid inhibition of mTORC1 activity, the tuberous sclerosis protein 2 (TSC2) tumor suppressor and the mTOR binding partner raptor. The chapter explores the molecular connections between AMPK and mTOR signaling pathways and examines the physiological processes in which AMPK regulation of mTOR is critical for growth or metabolic control. The AMP-activated protein kinase (AMPK) is a heterotrimeric kinase complex composed of a catalytic (α) subunit and two regulatory (β and γ) subunits.
Adenosine monophosphate-activated protein kinase (AMPK) is a conserved sensor of intracellular en... more Adenosine monophosphate-activated protein kinase (AMPK) is a conserved sensor of intracellular energy activated in response to low nutrient availability and environmental stress. In a screen for conserved substrates of AMPK, we identified ULK1 and ULK2, mammalian orthologs of the yeast protein kinase Atg1, which is required for autophagy. Genetic analysis of AMPK or ULK1 in mammalian liver and Caenorhabditis elegans revealed a requirement for these kinases in autophagy. In mammals, loss of AMPK or ULK1 resulted in aberrant accumulation of the autophagy adaptor p62 and defective mitophagy. Reconstitution of ULK1-deficient cells with a mutant ULK1 that cannot be phosphorylated by AMPK revealed that such phosphorylation is required for mitochondrial homeostasis and cell survival during starvation. These findings uncover a conserved biochemical mechanism coupling nutrient status with autophagy and cell survival.
Background: The appropriate control of mitotic entry and exit is reliant on a series of interlock... more Background: The appropriate control of mitotic entry and exit is reliant on a series of interlocking signaling events that coordinately drive the biological processes required for accurate cell division. Overlaid onto these signals that promote orchestrated cell division are checkpoints that ensure appropriate mitotic spindle formation, a lack of DNA damage, kinetochore attachment, and that each daughter cell has the appropriate complement of DNA. We recently discovered that AMP-activated protein kinase (AMPK) modulates the G2/M phase of cell cycle progression in part through its suppression of mammalian target of rapamycin (mTOR) signaling. AMPK directly phosphorylates the critical mTOR binding partner raptor inhibiting mTORC1 (mTOR-raptor rapamycin sensitive mTOR kinase complex 1). As mTOR has been previously tied to mitotic control, we examined further how raptor may contribute to this process.
AMPK is a highly conserved sensor of cellular energy status that is activated under conditions of... more AMPK is a highly conserved sensor of cellular energy status that is activated under conditions of low intracellular ATP. AMPK responds to energy stress by suppressing cell growth and biosynthetic processes, in part through its inhibition of the rapamycin-sensitive mTOR (mTORC1) pathway. AMPK phosphorylation of the TSC2 tumor suppressor contributes to suppression of mTORC1, however TSC2-deficient cells remain responsive to energy stress. Using a proteomic and bioinformatics approach, we sought to identify additional substrates of AMPK that mediate its effects on growth control. We report here that AMPK directly phosphorylates the mTOR binding partner raptor on two well-conserved serine residues, and this phosphorylation induces 14-3-3 binding to raptor. The phosphorylation of raptor by AMPK is required for the inhibition of mTORC1 and cell cycle arrest induced by energy stress. These findings uncover a novel conserved effector of AMPK that mediates its role as a metabolic checkpoint coordinating cell growth with energy status.
Resveratrol (3,5,49-trihydroxystilbene) extends the lifespan of diverse species including Sacchar... more Resveratrol (3,5,49-trihydroxystilbene) extends the lifespan of diverse species including Saccharomyces cerevisiae, Caenorhabditis elegans and Drosophila melanogaster. In these organisms, lifespan extension is dependent on Sir2, a conserved deacetylase proposed to underlie the beneficial effects of caloric restriction. Here we show that resveratrol shifts the physiology of middle-aged mice on a high-calorie diet towards that of mice on a standard diet and significantly increases their survival. Resveratrol produces changes associated with longer lifespan, including increased insulin sensitivity, reduced insulin-like growth factor-1 (IGF-I) levels, increased AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-c coactivator 1a (PGC-1a) activity, increased mitochondrial number, and improved motor function. Parametric analysis of gene set enrichment revealed that resveratrol opposed the effects of the high-calorie diet in 144 out of 153 significantly altered pathways. These data show that improving general health in mammals using small molecules is an attainable goal, and point to new approaches for treating obesity-related disorders and diseases of ageing.
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Papers by Dana Gwinn