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1.  Genomics: Drugs, diabetes and cancer 
Nature  2011;470(7334):338-339.
Variation in a genomic region that contains the cancer-a ssociated gene ATM affects a patient’s response to the diabetes drug metformin. Two experts discuss the implications for understanding diabetes and the link to cancer.
PMCID: PMC4237313  PMID: 21331030
2.  The autophagy initiating kinase ULK1 is regulated via opposing phosphorylation by AMPK and mTOR 
Autophagy  2011;7(6):645-646.
The serine/threonine kinase ULK1 is a mammalian homolog of Atg1, part of the Atg1 kinase complex, which is the most upstream component of the core autophagy machinery conserved from yeast to mammals. In budding yeast, activity of the Atg1 kinase complex is inhibited by TORC1 (target of rapamycin complex 1), but how the counterpart ULK1 complex in mammalian cells is regulated has been unknown. Our laboratories recently discovered that AMPK associates with, and directly phosphorylates, ULK1 on several sites and this modification is required for ULK1 activation after glucose deprivation. In contrast, when nutrients are plentiful, the mTORC1 complex phosphorylates ULK1, preventing its association and activation by AMPK. These studies have revealed a molecular mechanism of ULK1 regulation by nutrient signals via the actions of AMPK and mTORC1.
PMCID: PMC3359466  PMID: 21460621
autophagy; ULK1; AMPK; mTOR; 14-3-3
3.  A decade of molecular cell biology: achievements and challenges 
Nature Reviews Molecular Cell Biology celebrated its 10-year anniversary during this past year with a series of specially commissioned articles. To complement this, here we have asked researchers from across the field for their insights into how molecular cell biology research has evolved during this past decade, the key concepts that have emerged and the most promising interfaces that have developed. Their comments highlight the broad impact that particular advances have had, some of the basic understanding that we still require, and the collaborative approaches that will be essential for driving the field forward.
PMCID: PMC3282063  PMID: 21941276
4.  Metabolism and Cancer in La Jolla 
Cancer research  2010;70(10):3864-3869.
PMCID: PMC2873170  PMID: 20442287
5.  LKB1 and AMPK control of mTOR signalling and growth 
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 ATP following stresses such as nutrient deprivation or hypoxia. In the past five years, work from a large number of laboratories has revealed that one of the major downstream signaling pathways regulated by AMPK is the mammalian target-of-rapamycin (mTOR pathway). Interestingly, like AMPK, the mTOR serine/threonine kinase plays key roles not only in growth control and cell proliferation but also in metabolism. Recent work has revealed that 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. Many details of the molecular mechanism by which AMPK inhibits mTORC1 signaling have also been decoded in the past 5 years. AMPK directly phosphorylates at least two proteins to induce rapid suppression of mTORC1 activity, the TSC2 tumor suppressor and the critical mTORC1 binding subunit raptor. Here we explore the molecular connections between AMPK and mTOR signaling pathways and examine the physiological processes in which AMPK regulation of mTOR is critical for growth or metabolic control. The functional conservation of AMPK and TOR in all eukaryotes, and the sequence conservation around the AMPK phosphorylation sites in raptor across all eukaryotes examined suggest that this represents a fundamental cell growth module connecting nutrient status to the cell growth machinery. These findings have broad implications for the control of cell growth by nutrients in a number of cellular and organismal contexts.
PMCID: PMC2760308  PMID: 19245654
LKB1; AMPK; mTOR; raptor; TSC2; metabolism; checkpoint
6.  The LKB1-AMPK pathway: metabolism and growth control in tumor suppression 
Nature reviews. Cancer  2009;9(8):563-575.
In the past decade, studies of the human tumor suppressor LKB1 have uncovered a novel signaling pathway that links cell metabolism to growth control and cell polarity. LKB1 encodes a serine/threonine kinase that directly phosphorylates and activates AMPK, a central metabolic sensor. AMPK regulates lipid, cholesterol and glucose metabolism in specialized metabolic tissues such as liver, muscle, and adipose, a function that has made it a key therapeutic target in patients with diabetes. The connection of AMPK with several tumor suppressors suggests that therapeutic manipulation of this pathway with established diabetes drugs warrants further investigation in patients with cancer.
PMCID: PMC2756045  PMID: 19629071
7.  AMPK phosphorylation of raptor mediates a metabolic checkpoint 
Molecular cell  2008;30(2):214-226.
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.
PMCID: PMC2674027  PMID: 18439900
8.  mTOR Complex 2 Controls Glycolytic Metabolism in Glioblastoma through FoxO Acetylation and Upregulation of c-Myc 
Cell metabolism  2013;18(5):10.1016/j.cmet.2013.09.013.
Aerobic glycolysis (Warburg effect) is a core hallmark of cancer, but the molecular mechanisms underlying it remain unclear. Here, we identify an unexpected central role for mTORC2 in cancer metabolic reprogramming where it controls glycolytic metabolism by ultimately regulating the cellular level of c-Myc. We show that mTORC2 promotes inactivating phosphorylation of class IIa histone deacetylases that leads to the acetylation of FoxO1 and FoxO3, and this in turn releases c-Myc from a suppressive miR-34c-dependent network. These central features of activated mTORC2 signaling, acetylated FoxO and c-Myc levels are highly inter-correlated in clinical samples, and with shorter survival of GBM patients. These results identify a specific, Akt-independent, role for mTORC2 in regulating glycolytic metabolism in cancer.
PMCID: PMC3840163  PMID: 24140020
9.  Metabolic and Functional Genomic Studies Identify Deoxythymidylate Kinase as a target in LKB1 Mutant Lung Cancer 
Cancer discovery  2013;3(8):870-879.
The LKB1/STK11 tumor suppressor encodes a serine/threonine kinase which coordinates cell growth, polarity, motility, and metabolism. In non-small cell lung cancer, LKB1 is somatically inactivated in 25-30% of cases, often concurrently with activating KRAS mutation. Here, we employed an integrative approach to define novel therapeutic targets in KRAS-driven LKB1 mutant lung cancers. High-throughput RNAi screens in lung cancer cell lines from genetically engineered mouse models driven by activated KRAS with or without coincident Lkb1 deletion led to the identification of Dtymk, encoding deoxythymidylate kinase which catalyzes dTTP biosynthesis, as synthetically lethal with Lkb1 deficiency in mouse and human lung cancer lines. Global metabolite profiling demonstrated that Lkb1-null cells had striking decreases in multiple nucleotide metabolites as compared to the Lkb1-wt cells. Thus, LKB1 mutant lung cancers have deficits in nucleotide metabolism conferring hypersensitivity to DTYMK inhibition, suggesting that DTYMK is a potential therapeutic target in this aggressive subset of tumors.
PMCID: PMC3753578  PMID: 23715154
LKB1; KRAS; DTYMK; CHEK1; NSCLC; GEMM-derived cell line; genome wide RNAi screen; metabolic profiling
11.  LKB1 inactivation dictates therapeutic response of non-small cell lung cancer to the metabolism drug phenformin 
Cancer cell  2013;23(2):143-158.
The LKB1 (also called STK11) tumor suppressor is mutationally inactivated in ~20% of non-small cell lung cancers (NSCLC). LKB1 is the major upstream kinase activating the energy-sensing kinase AMPK, making LKB1-deficient cells unable to appropriately sense metabolic stress. We tested the therapeutic potential of metabolic drugs in NSCLC and identified phenformin, a mitochondrial inhibitor and analog of the diabetes therapeutic metformin, as selectively inducing apoptosis in LKB1-deficient NSCLC cells. Therapeutic trials in Kras-dependent mouse models of NSCLC revealed that tumors with Kras and Lkb1 mutations, but not those with Kras and p53 mutations showed selective response to phenformin as a single agent, resulting in prolonged survival. This study suggests phenformin as a cancer metabolism-based therapeutic to selectively target LKB1-deficient tumors.
PMCID: PMC3579627  PMID: 23352126
12.  Metabolic Reprogramming by Class I and II Histone Deacetylases 
Accumulating evidence suggests that protein acetylation plays a major regulatory role in many facets of transcriptional control of metabolism. The enzymes that catalyze the addition and removal of acetyl moieties are the Histone Acetyl Transferases (HATs) and Histone Deacetylases (HDACs), respectively. A number of recent studies have uncovered novel mechanisms and contexts in which different HDACs play critical roles in metabolic control. Understanding the role of Class I and II HDACs in different metabolic programs during development, as well as in the physiology and pathology of the adult organism, will lead to novel therapeutics for metabolic disease. Here, we review the current understanding of how Class I and Class II HDACs contribute to metabolic control.
PMCID: PMC3532556  PMID: 23062770
HDAC; HAT; lipogenesis; gluconeogenesis; AMPK; HDAC3
13.  Reduced cell proliferation by IKK2 depletion in a mouse lung cancer model 
Nature cell biology  2012;14(3):257-265.
Lung cancer is one of the leading cancer malignancies with a five-year survival rate of only ~15%. We have developed a lentiviral vector mediated mouse model which allows generation of non-small cell lung cancer from less than one hundred alveolar epithelial cells, and investigated the role of IKK2 and NF-κB in lung cancer development. IKK2 depletion in tumour cells significantly attenuated tumour proliferation and significantly prolonged mouse survival. We identified Timp-1, one of the NF-κB target genes, as a key mediator for tumour growth. Activation of Erk signalling pathway and cell proliferation requires Timp-1 and its receptor CD63. Knockdown of either IKK2 or Timp-1 by shRNAs reduced tumour growth in both xenograft and lentiviral models. Our results, thus suggest the possible application of IKK2 and Timp-1 inhibitors in treating lung cancer.
PMCID: PMC3290728  PMID: 22327365
14.  Characterization of a Hormone Dependent Module Regulating Energy Balance 
Cell  2011;145(4):596-606.
Under fasting conditions, metazoans maintain energy balance by shifting from glucose to fat burning. In the fasted state, SIRT1 promotes catabolic gene expression by deacetylating the forkhead factor FOXO in response to stress and nutrient deprivation. The mechanisms by which hormonal signals regulate FOXO deacetylation remain unclear, however. We identified a hormone-dependent module, consisting of the Ser/Thr kinase SIK3 and the class IIa deacetylase HDAC4, which regulates FOXO activity in Drosophila. During feeding, HDAC4 is phosphorylated and sequestered in the cytoplasm by SIK3, whose activity is upregulated in response to insulin. SIK3 is inactivated during fasting, leading to the de-phosphorylation and nuclear translocation of HDAC4, and to FOXO deacetylation. SIK3 mutant flies are starvation-sensitive, reflecting FOXO-dependent increases in lipolysis that deplete triglyceride stores; reducing HDAC4 expression restored lipid accumulation. Our results reveal a hormone-regulated pathway that functions in parallel with the nutrient-sensing SIRT1 pathway to maintain energy balance.
PMCID: PMC3129781  PMID: 21565616
15.  AMPK Phosphorylates and Inhibits SREBP Activity to Attenuate Hepatic Steatosis and Atherosclerosis in Diet-induced Insulin Resistant Mice 
Cell metabolism  2011;13(4):376-388.
AMPK has emerged as a critical mechanism for salutary effects of polyphenols on lipid metabolic disorders in type 1 and type 2 diabetes. We demonstrate that AMPK interacts with and directly phosphorylates sterol regulatory element binding proteins (SREBP-1c and −2). Ser372 phosphorylation of SREBP-1c by AMPK is sufficient and necessary for inhibition of proteolytic processing and transcriptional activity of SREBP-1c in response to polyphenols and metformin. AMPK stimulates Ser372 phosphorylation, suppresses SREBP-1c cleavage and nuclear translocation, and represses SREBP-1c target gene expression in hepatocytes exposed to high glucose, leading to reduced lipogenesis and lipid accumulation. Hepatic activation of AMPK by the synthetic polyphenol S17834 protects against hepatic steatosis, hyperlipidemia, and accelerated atherosclerosis in diet-induced insulin resistant LDL receptor deficient mice in part through phosphorylation of SREBP-1c Ser372 and suppression of SREBP-1c and −2-dependent lipogenesis. AMPK-dependent phosphorylation of SREBP may offer novel therapeutic strategies to combat insulin resistance, dyslipidemia, and atherosclerosis.
PMCID: PMC3086578  PMID: 21459323
16.  The AMP-activated protein kinase (AMPK) signaling pathway coordinates cell growth, autophagy, & metabolism 
Nature cell biology  2011;13(9):1016-1023.
One of the central regulators of cellular and organismal metabolism in eukaryotes is the AMP-activated protein kinase (AMPK), which is activated when intracellular ATP levels lower. AMPK plays critical roles in regulating growth and reprogramming metabolism, and recently has been connected to cellular processes including autophagy and cell polarity. We review here a number of recent breakthroughs in the mechanistic understanding of AMPK function, focusing on a number of new identified downstream effectors of AMPK.
PMCID: PMC3249400  PMID: 21892142
17.  Decoding key nodes in the metabolism of cancer cells: sugar & spice and all things nice 
In the past 5 years, a convergence of studies has resulted in a broad appreciation in the cancer research community that reprogramming of cellular metabolism may be more central to cancer than appreciated in the past 30 years. The re-emergence of cancer metabolism stems in part from discoveries that a number of common oncogenes and tumor suppressor genes more directly control cell metabolism than previously thought. In addition, a number of what would previously have been called “card-carrying” metabolic enzymes have been identified as human tumor suppressors or oncogenes, causally mutated in a variety of human cancers. This growing appreciation of the role of altered cell metabolism has led to further investigation into the rate-limiting proteins involved in different aspects of the unique metabolism of tumor cells. Targeting cancer metabolism with drugs requires a therapeutic window in which tumor cells, compared to normal tissues, have a greater dependence on specific metabolic enzymes. Themes that have emerged in the past decade of developing oncogene-targeted cancer therapeutics suggest that tumors with distinct oncogenic lesions are likely to require drugs that target distinct metabolic pathways. Ultimately, the hope is that detailed knowledge of oncogene and tumor suppressor gene functions and their effects on metabolism will lead to drug combinations that will be far more effective in treating cancers.
PMCID: PMC3255319  PMID: 22242042
18.  Class IIa Histone Deacetylases are Hormone-activated regulators of FOXO and Mammalian Glucose Homeostasis 
Cell  2011;145(4):607-621.
Class IIa histone deacetylases (HDACs) are signal-dependent modulators of transcription with established roles in muscle differentiation and neuronal survival. We show here that in liver, Class IIa HDACs (HDAC4, 5, and 7) are phosphorylated and excluded from the nucleus by AMPK family kinases. In response to the fasting hormone glucagon, Class IIa HDACs are rapidly dephosphorylated and translocated to the nucleus where they associate with the promoters of gluconeogenic enzymes such as G6Pase. In turn, HDAC4/5 recruit HDAC3, which results in the acute transcriptional induction of these genes via deacetylation and activation of Foxo family transcription factors. Loss of Class IIa HDACs in murine liver results in inhibition of FOXO target genes and lowers blood glucose, resulting in increased glycogen storage. Finally, suppression of Class IIa HDACs in mouse models of Type 2 Diabetes ameliorates hyperglycemia, suggesting that inhibitors of Class I/II HDACs may be potential therapeutics for metabolic syndrome.
PMCID: PMC3117637  PMID: 21565617
19.  Lifespan extension induced by AMPK and calcineurin is mediated by CRTC-1 and CREB 
Nature  2011;470(7334):404-408.
Activating AMPK or inactivating calcineurin slows ageing in Caenorhabditis elegans1,2 and both have been implicated as therapeutic targets for age-related pathology in mammals3–5. However, the direct targets that mediate their effects on longevity remain unclear. In mammals, CREB-regulated transcriptional coactivators (CRTCs)6 are a family of cofactors involved in diverse physiological processes including energy homeostasis7–9, cancer10 and endoplasmic reticulum stress11. Here we show that both AMPK and calcineurin modulate longevity exclusively through post-translational modification of CRTC-1, the sole C. elegans CRTC. We demonstrate that CRTC-1 is a direct AMPK target, and interacts with the CREB homologue-1 (CRH-1) transcription factor in vivo. The pro-longevity effects of activating AMPK or deactivating calcineurin decrease CRTC-1 and CRH-1 activity and induce transcriptional responses similar to those of CRH-1 null worms. Downregulation of crtc-1 increases lifespan in a crh-1-dependent manner and directly reducing crh-1 expression increases longevity, substantiating a role for CRTCs and CREB in ageing. Together, these findings indicate a novel role for CRTCs and CREB in determining lifespan downstream of AMPK and calcineurin, and illustrate the molecular mechanisms by which an evolutionarily conserved pathway responds to low energy to increase longevity.
PMCID: PMC3098900  PMID: 21331044
20.  Phosphorylation of ULK1 (hATG1) by AMP-activated protein kinase connects energy sensing to mitophagy 
Science (New York, N.Y.)  2010;331(6016):456-461.
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 C. 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 following starvation. These findings uncover a conserved biochemical mechanism coupling nutrient status with autophagy and cell survival.
PMCID: PMC3030664  PMID: 21205641
21.  The Kinase LKB1 Mediates Glucose Homeostasis in Liver and Therapeutic Effects of Metformin 
Science (New York, N.Y.)  2005;310(5754):1642-1646.
The Peutz-Jegher syndrome tumor-suppressor gene encodes a protein-threonine kinase, LKB1, which phosphorylates and activates AMPK [adenosine monophosphate (AMP)–activated protein kinase]. The deletion of LKB1 in the liver of adult mice resulted in a nearly complete loss of AMPK activity. Loss of LKB1 function resulted in hyperglycemia with increased gluconeogenic and lipogenic gene expression. In LKB1-deficient livers, TORC2, a transcriptional coactivator of CREB (cAMP response element–binding protein), was dephosphorylated and entered the nucleus, driving the expression of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), which in turn drives gluconeogenesis. Adenoviral small hairpin RNA (shRNA) for TORC2 reduced PGC-1α expression and normalized blood glucose levels in mice with deleted liver LKB1, indicating that TORC2 is a critical target of LKB1/AMPK signals in the regulation of gluconeogenesis. Finally, we show that metformin, one of the most widely prescribed type 2 diabetes therapeutics, requires LKB1 in the liver to lower blood glucose levels.
PMCID: PMC3074427  PMID: 16308421
22.  Raptor is Phosphorylated by cdc2 during Mitosis 
PLoS ONE  2010;5(2):e9197.
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.
Methodology/Principal Findings
We have discovered that raptor becomes highly phosphorylated in cells in mitosis. Utilizing tandem mass spectrometry, we identified a number of novel phosphorylation sites in raptor, and using phospho-specific antibodies demonstrated that raptor becomes phosphorylated on phospho-serine/threonine-proline sites in mitosis. A combination of site-directed mutagenesis in a tagged raptor cDNA and analysis with a series of new phospho-specific antibodies generated against different sites in raptor revealed that Serine 696 and Threonine 706 represent two key sites in raptor phosphorylated in mitosis. We demonstrate that the mitotic cyclin-dependent kinase cdc2/CDK1 is the kinase responsible for phosphorylating these sites, and its mitotic partner Cyclin B efficiently coimmunoprecipitates with raptor in mitotic cells.
This study demonstrates that the key mTOR binding partner raptor is directly phosphorylated during mitosis by cdc2. This reinforces previous studies suggesting that mTOR activity is highly regulated and important for mitotic progression, and points to a direct modulation of the mTORC1 complex during mitosis.
PMCID: PMC2820552  PMID: 20169205
23.  AMPK Regulates the Circadian Clock by Cryptochrome Phosphorylation and Degradation 
Science (New York, N.Y.)  2009;326(5951):437-440.
Circadian clocks coordinate behavioral and physiological processes with daily light-dark cycles by driving rhythmic transcription of thousands of genes. Whereas the master clock in the brain is set by light, pacemakers in peripheral organs, such as the liver, are reset by food availability, although the setting, or “entrainment,” mechanisms remain mysterious. Studying mouse fibroblasts, we demonstrated that the nutrient-responsive adenosine monophosphate–activated protein kinase (AMPK) phosphorylates and destabilizes the clock component cryptochrome 1 (CRY1). In mouse livers, AMPK activity and nuclear localization were rhythmic and inversely correlated with CRY1 nuclear protein abundance. Stimulation of AMPK destabilized cryptochromes and altered circadian rhythms, and mice in which the AMPK pathway was genetically disrupted showed alterations in peripheral clocks. Thus, phosphorylation by AMPK enables cryptochrome to transduce nutrient signals to circadian clocks in mammalian peripheral organs.
PMCID: PMC2819106  PMID: 19833968
24.  mTOR signaling: RAG GTPases transmit the amino acid signal 
Trends in biochemical sciences  2008;33(12):565-568.
mTOR (mammalian target of rapamycin) is a highly conserved nutrient-responsive regulator of cell growth that is found in all eukaryotes. The mechanism by which amino acids signal to mTOR has remained one of the largest outstanding questions in the field. Two recent complimentary studies provide compelling evidence that the Rag family of small GTPases is both necessary and sufficient to transmit a positive signal from amino acids to mTOR.
PMCID: PMC2677387  PMID: 18929489
25.  LKB1: cancer, polarity, metabolism, and now fertility 
The Biochemical journal  2008;416(1):e1-e3.
The LKB1 serine/threonine kinase is a tumor suppressor responsible for the inherited familial cancer disorder Peutz-Jeghers syndrome and is inactivated in a large percentage of human lung cancers. LKB1 acts a master kinase, directly phosphorylating and activating a family of 14 AMPK-related kinases which control cell metabolism, cell growth, and cell polarity. In this issue of Biochemical Journal, Hardie and colleagues discover an alternative splice form of LKB1 that alters the C-terminus of the protein containing a few known sites of post-translational regulation. Though widely expressed, the short isoform (LKB1s) is the sole splice isoform expressed in testes and its expression peaks at the time of spermatid maturation. Male mice lacking the LKB1s isoform have dramatic defects in spermatozoa resulting in sterility.
PMCID: PMC2676712  PMID: 18939946
LKB1; AMPK; kinase; alternative splicing; spermatogenesis

Results 1-25 (27)