Cellular Energy Levels Research Articles

Adenylate Charge Regulates Sensor Kinase CheS3 To Control Cyst Formation in Rhodospirillum centenum.

ABSTRACTRhodospirillum centenum forms metabolically dormant cysts under unfavorable growth conditions such as desiccation or nutrient starvation. The development of cysts is tightly regulated and involves a cyst-repressing chemotaxis-like signal transduction pathway called the Che3 signaling cascade. The Che3 cascade is comprised of a methyl chemoreceptor (MCP3), receptor-methylating/demethylating proteins CheB3 and CheR3, two CheW3 linker proteins, a CheA3-CheY hybrid histidine kinase, and a single-domain response regulator, CheY3. In addition to Che-like components, the Che3 cascade also contains a second hybrid histidine kinase, CheS3. Recent biochemical and genetic studies show that CheA3 does not serve as a phosphor donor for CheY3; instead, CheA3 inhibits a CheS3→CheY3 two-component system by phosphorylating an inhibitory receiver domain of CheS3. In this study, we show that in addition to phosphorylation by CheA3, the phosphorylation state of CheS3 is also regulated by the cellular energy level as quantified by the molar ratio of ATP/(ATP + ADP). A 35% decrease in cellular energy is shown to occur in vivo upon a nutrient downshift that gives rise to cyst formation. When this energy decline is replicated in vitro, the phosphorylation level of CheS3 is reduced by ~75%. Finally, we also show that ADP-mediated reduction of CheS3 phosphorylation is a consequence of ADP enhancing autodephosphorylation of CheS3.

Negative regulation of the LKB1/AMPK pathway by ERK in human acute myeloid leukemia cells

Adenosine monophosphate-activated protein kinase (AMPK) is a sensor for cellular energy status. When the cellular energy level is decreased, AMPK is activated and functions to suppress energy-consuming processes, including protein synthesis. Recently, AMPK has received attention as an attractive molecular target for cancer therapy. Several studies have revealed that the activation of AMPK by chemical stimulators, such as metformin, induces apoptosis in a variety of hematologic malignant cells. From another perspective, these results suggest that the function of AMPK is impaired in hematologic tumor cells. However, the precise mechanisms by which this impairment occurs are not well understood. In melanoma cells, oncogenic BRAF constitutively activates the extracellular signal-regulated kinase (ERK) pathway and phosphorylates liver kinase B1, an upstream activator of 5' adenosine monophosphate-activated protein kinase (AMPK), resulting in the inactivation of liver kinase B1 and AMPK. In this study, we analyzed whether ERK is involved in the suppression of AMPK activity using established and primary human leukemia cells. We found an inverse correlation between the intensity of ERK activity and the degree of AMPK activation after stimulation with either glucose deprivation or metformin. We also found that the inhibition of ERK activity by U0126 restored AMPK activation after metformin treatment. Furthermore, a combined treatment with metformin and U0126 enhanced the antileukemic activity of metformin. Importantly, metformin induced ERK activation by suppressing the protein levels of dual specificity phosphatase 6, a negative regulator of ERK. This crosstalk between AMPK and ERK could diminish the antileukemic activity of metformin. Taken together, our present observations suggest a novel therapeutic strategy for improving the efficacy of metformin in treating leukemia.

Computational analysis of an autophagy/translation switch based on mutual inhibition of MTORC1 and ULK1.

We constructed a mechanistic, computational model for regulation of (macro)autophagy and protein synthesis (at the level of translation). The model was formulated to study the system-level consequences of interactions among the following proteins: two key components of MTOR complex 1 (MTORC1), namely the protein kinase MTOR (mechanistic target of rapamycin) and the scaffold protein RPTOR; the autophagy-initiating protein kinase ULK1; and the multimeric energy-sensing AMP-activated protein kinase (AMPK). Inputs of the model include intrinsic AMPK kinase activity, which is taken as an adjustable surrogate parameter for cellular energy level or AMP:ATP ratio, and rapamycin dose, which controls MTORC1 activity. Outputs of the model include the phosphorylation level of the translational repressor EIF4EBP1, a substrate of MTORC1, and the phosphorylation level of AMBRA1 (activating molecule in BECN1-regulated autophagy), a substrate of ULK1 critical for autophagosome formation. The model incorporates reciprocal regulation of mTORC1 and ULK1 by AMPK, mutual inhibition of MTORC1 and ULK1, and ULK1-mediated negative feedback regulation of AMPK. Through analysis of the model, we find that these processes may be responsible, depending on conditions, for graded responses to stress inputs, for bistable switching between autophagy and protein synthesis, or relaxation oscillations, comprising alternating periods of autophagy and protein synthesis. A sensitivity analysis indicates that the prediction of oscillatory behavior is robust to changes of the parameter values of the model. The model provides testable predictions about the behavior of the AMPK-MTORC1-ULK1 network, which plays a central role in maintaining cellular energy and nutrient homeostasis.

Activation of AMP-activated protein kinase in response to temperature elevation shows seasonal variation in the zebra mussel, Dreissena polymorpha

Global climate change is affecting ectothermic species, and a variety of studies are needed on thermal tolerances, especially from cellular and physiological perspectives. This study utilized AMP-activated protein kinase (AMPK), a key regulator of cellular energy levels, to examine the effects of high water temperatures on zebra mussel (Dreissena polymorpha) physiology. During heating, AMPK activity increased as water temperature increased to a point, and maximum AMPK activity was detected at high, but sublethal, water temperatures. This pattern varied with season, suggesting that cellular mechanisms of seasonal thermal acclimatization affect basic metabolic processes during sublethal heat stress. There was a greater seasonal variation in the water temperature at which maximum AMPK activity was measured than in lethal water temperature. Furthermore, baseline AMPK activity varied significantly across seasons, most likely reflecting altered metabolic states during times of growth and reproduction. In addition, when summer-collected mussels were lab-acclimated to winter and spring water temperatures, patterns of heat stress mirrored those of field-collected animals. These data suggest that water temperature is the main driver of the seasonal variation in physiology. This study concluded that AMPK activity, which reflects changes in energy supply and demand during heat stress, can serve as a sensitive and early indicator of temperature stress in mussels.

Betaine is a positive regulator of mitochondrial respiration

Betaine protects cells from environmental stress and serves as a methyl donor in several biochemical pathways. It reduces cardiovascular disease risk and protects liver cells from alcoholic liver damage and nonalcoholic steatohepatitis. Its pretreatment can rescue cells exposed to toxins such as rotenone, chloroform, and LiCl. Furthermore, it has been suggested that betaine can suppress cancer cell growth in vivo and in vitro. Mitochondrial electron transport chain (ETC) complexes generate the mitochondrial membrane potential, which is essential to produce cellular energy, ATP. Reduced mitochondrial respiration and energy status have been found in many human pathological conditions including aging, cancer, and neurodegenerative disease. In this study we investigated whether betaine directly targets mitochondria. We show that betaine treatment leads to an upregulation of mitochondrial respiration and cytochrome c oxidase activity in H2.35 cells, the proposed rate limiting enzyme of ETC in vivo. Following treatment, the mitochondrial membrane potential was increased and cellular energy levels were elevated. We propose that the anti-proliferative effects of betaine on cancer cells might be due to enhanced mitochondrial function contributing to a reversal of the Warburg effect.

Increased sucrose levels mediate selective mRNA translation in Arabidopsis.

BackgroundProtein synthesis is a highly energy demanding process and is regulated according to cellular energy levels. Light and sugar availability affect mRNA translation in plant cells but the specific roles of these factors remain unclear. In this study, sucrose was applied to Arabidopsis seedlings kept in the light or in the dark, in order to distinguish sucrose and light effects on transcription and translation. These were studied using microarray analysis of steady-state mRNA and mRNA bound to translating ribosomes.ResultsSteady-state mRNA levels were affected differently by sucrose in the light and in the dark but general translation increased to a similar extent in both conditions. For a majority of the transcripts changes of the transcript levels were followed by changes in polysomal mRNA levels. However, for 243 mRNAs, a change in polysomal occupancy (defined as polysomal levels related to steady-state levels of the mRNA) was observed after sucrose treatment in the light, but not in the dark condition. Many of these mRNAs are annotated as encoding ribosomal proteins, supporting specific translational regulation of this group of transcripts. Unexpectedly, the numbers of ribosomes bound to each mRNA decreased for mRNAs with increased polysomal occupancy.ConclusionsOur results suggest that sucrose regulate translation of these 243 mRNAs specifically in the light, through a novel regulatory mechanism. Our data shows that increased polysomal occupancy is not necessarily leading to more ribosomes per transcript, suggesting a mechanism of translational induction not solely dependent on increased translation initiation rates.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0306-3) contains supplementary material, which is available to authorized users.

Abstract 329: Rottlerin induced autophagy by targeting multiple sites that leads to the apoptosis in cancer stem cells

Abstract Autophagy plays an important role in maintaining cellular process that controls cells in a normal homeostatic state by recycling nutrients to maintain cellular energy levels for cell survival via degradation of aggregated proteins and damaged organelles. Autophagy is associated with the several catabolic and pathological processes, including cancer. Cancer is a multifaceted disorder involving the perturbation of several different pathways that regulate cell proliferation, cell differentiation, and cell survival. However, persistent activation of autophagy can lead to excessive depletion of cellular organelles and essential proteins, leading to apoptosis in cancer stem cells. As such, inducing cell death through this autophagic mechanism could be an alternative approach to the treatment of cancers. Recently, we have identified a novel autophagic inducer, rottlerin, from a medicinal plant (Mallotus philippinensis) that induces autophagy in various types of cancer stem cells through the formation of autophagosomes as measured by electron microscopy and GFP-LC3 puncta formation. By computational virtual docking analysis, rottlerin was shown to interact directly with the protein kinase-C delta, beclin-1, and autophagy related proteins. By biochemical assays, rottlerin was shown to increase autophagy induction through the activation of the kinase-AMP-activated protein kinase-mammalian target of rapamycin pathway. In addition, rottlerin treatment causes the induction of apoptosis, which was associated with the suppression of phosphorylated Akt and mTOR, upregulation of phosphorylated kinase-AMP, downregulation of Bcl-2, Bcl-XL, XIAP, and cIAP-1. Rottlerin also proved to be a potent autophagy inducer in cancer stem cells, by knocking down beclin-1, protein kinase-C delta, or Agt7. These results provide a detailed understanding of the mechanism of action of rottlerin, as a novel autophagic inducer by targeting multiple sites to induce autophagy, which leads to the apoptosis in cancer stem cells. Rottlerin has the potential of being developed into an anti-cancer agent by targeting cancer stem cells. Citation Format: Dhruv Kumar. Rottlerin induced autophagy by targeting multiple sites that leads to the apoptosis in cancer stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 329. doi:10.1158/1538-7445.AM2014-329

Xanthene derivatives increase glucose utilization through activation of LKB1-dependent AMP-activated protein kinase.

5′ AMP-activated protein kinase (AMPK) is a highly conserved serine-threonine kinase that regulates energy expenditure by activating catabolic metabolism and suppressing anabolic pathways to increase cellular energy levels. Therefore AMPK activators are considered to be drug targets for treatment of metabolic diseases such as diabetes mellitus. To identify novel AMPK activators, we screened xanthene derivatives. We determined that the AMPK activators 9H-xanthene-9-carboxylic acid {2,2,2-trichloro-1-[3-(3-nitro-phenyl)-thioureido]-ethyl}-amide (Xn) and 9H-xanthene-9-carboxylic acid {2,2,2-trichloro-1-[3-(3-cyano-phenyl)-thioureido]-ethyl}-amide (Xc) elevated glucose uptake in L6 myotubes by stimulating translocation of glucose transporter type 4 (GLUT4). Treatment with the chemical AMPK inhibitor compound C and infection with dominant-negative AMPKa2-virus inhibited AMPK phosphorylation and glucose uptake in myotubes induced by either Xn or Xc. Of the two major upstream kinases of AMPK, we found that Xn and Xc showed LKB1 dependency by knockdown of STK11, an ortholog of human LKB1. Single intravenous administration of Xn and Xc to high-fat diet-induced diabetic mice stimulated AMPK phosphorylation of skeletal muscle and improved glucose tolerance. Taken together, these results suggest that Xn and Xc regulate glucose homeostasis through LKB1-dependent AMPK activation and that the compounds are potential candidate drugs for the treatment of type 2 diabetes mellitus.

Activation of 5' adenosine monophosphate-activated protein kinase blocks cumulus cell expansion through inhibition of protein synthesis during in vitro maturation in Swine.

The serine/threonine kinase 5' adenosine monophosphate-activated protein kinase (AMPK), a heterotrimeric protein known as a metabolic switch, is involved in oocyte nuclear maturation in mice, cattle, and swine. The present study analyzed AMPK activation in cumulus cell expansion during in vitro maturation (IVM) of porcine cumulus-oocyte complexes (COC). 5-Aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR) is a well-known activator of AMPK. It inhibited oocyte meiotic resumption in COC. Moreover, cumulus cell expansion did not occur in the presence of AICAR, demonstrating its marked impact on cumulus cells. Activation of AMPK was supported by AICAR-mediated phosphorylation of alpha AMPK subunits. Furthermore, the presence of AICAR increased glucose uptake, a classical response to activation of this metabolic switch in response to depleted cellular energy levels. Neither nuclear maturation nor cumulus expansion was reversed by glucosamine, an alternative substrate in hyaluronic acid synthesis, through the hexosamine biosynthetic pathway, which ruled out possible depletion of substrates. Both increased gap junction communication and phosphodiesterase activity in COC are dependent on protein synthesis during the initial hours of IVM; however, both were inhibited in the presence of AICAR, which supports the finding that activation of AMPK by AICAR mediated inhibition of protein synthesis. Moreover, this protein synthesis inhibition was equivalent to that of the well-known protein synthesis inhibitor cycloheximide, as observed on cumulus expansion and protein concentration. Finally, the phosphorylation level of selected kinases was investigated. The pattern of raptor phosphorylation is supportive of activation of AMPK-mediated inhibition of protein synthesis. In conclusion, AICAR-mediated AMPK activation in porcine COC inhibited cumulus cell expansion and protein synthesis. These results bring new considerations to the importance of this kinase in ovarian physiology and to the development of new oocyte culture medium.

Energy, the driving force behind good and ill health.

Energy, the driving force behind good and ill health.

AMPK activation: a therapeutic target for type 2 diabetes?

Type 2 diabetes (T2D) is a metabolic disease characterized by insulin resistance, β-cell dysfunction, and elevated hepatic glucose output. Over 350 million people worldwide have T2D, and the International Diabetes Federation projects that this number will increase to nearly 600 million by 2035. There is a great need for more effective treatments for maintaining glucose homeostasis and improving insulin sensitivity. AMP-activated protein kinase (AMPK) is an evolutionarily conserved serine/threonine kinase whose activation elicits insulin-sensitizing effects, making it an ideal therapeutic target for T2D. AMPK is an energy-sensing enzyme that is activated when cellular energy levels are low, and it signals to stimulate glucose uptake in skeletal muscles, fatty acid oxidation in adipose (and other) tissues, and reduces hepatic glucose production. There is substantial evidence suggesting that AMPK is dysregulated in animals and humans with metabolic syndrome or T2D, and that AMPK activation (physiological or pharmacological) can improve insulin sensitivity and metabolic health. Numerous pharmacological agents, natural compounds, and hormones are known to activate AMPK, either directly or indirectly – some of which (for example, metformin and thiazolidinediones) are currently used to treat T2D. This paper will review the regulation of the AMPK pathway and its role in T2D, some of the known AMPK activators and their mechanisms of action, and the potential for future improvements in targeting AMPK for the treatment of T2D.

Mechanisms and consequences of hepatic regulation of mTORC1 by metformin

Background In mammals, the ability to sense and respond to both intracellular and extracellular nutrient levels requires the integration and cooperation of multiple complex metabolic regulatory networks. Key among these are the mTOR and AMPK signaling pathways, which are activated in response to increased or decreased cellular energy levels, respectively. These pathways control cell growth, proliferation, metabolism and autophagy, and their dysregulation has been associated with both cancer and metabolic disease. The anti-diabetes drug metformin has been shown to function, at least in part, through activation of AMPK, however it also inhibits mTORC1 signaling, suggesting a potential role for mTORC1 inhibition in the beneficial effects of metformin. AMPK can suppress mTORC1 through activation of the TSC complex, an inhibitor of mTORC1, or inhibition of Raptor, an essential component of mTORC1. The liver plays a critical role in the maintenance of systemic metabolic homeostasis, and is thought to be the major target organ for the physiological effects of metformin, thus we are investigating the hepatic regulation of mTORC1 by metformin. Materials and methods The effects of metformin on mTORC1 were assessed in livers and primary hepatocytes from LTSC1KO mice that have conditional loss of TSC1 in the liver. mTORC1 activity was determined by analysis of the phosphorylation state of its targets, and the rate of protein synthesis using 35S-methionine incorporation. Physiological effects of metformin were determined using glucose and metformin tolerance tests. Results

Perspectives of the AMP-activated kinase (AMPK) signalling pathway in thyroid cancer.

Approximately 90% of non-medullary thyroid malignancies originate from the follicular cell and are classified as papillary or follicular (well-differentiated) thyroid carcinomas, showing an overall favourable prognosis. However, recurrence or persistence of the disease occurs in some cases associated with the presence of loco-regional or distant metastatic lesions that generally become resistant to radioiodine therapy, while glucose uptake and metabolism are increased. Recent advances in the field of tumor progression have shown that CTC (circulating tumour cells) are metabolic and genetically heterogeneous. There is now special interest in unravelling the mechanisms that allow the reminiscence of dormant tumour lesions that might be related to late disease progression and increased risk of recurrence. AMPK (AMP-activated protein kinase) is activated by the depletion in cellular energy levels and allows adaptive changes in cell metabolism that are fundamental for cell survival in a stressful environment; nevertheless, the activation of this kinase also decreases cell proliferation rate and induces tumour cell apoptosis. In the thyroid field, AMPK emerged as a novel important intracellular pathway, since it regulates both iodide and glucose uptakes in normal thyroid cells. Furthermore, it has recently been demonstrated that the AMPK pathway is highly activated in papillary thyroid carcinomas, although the clinical significance of these findings remains elusive. Herein we review the current knowledge about the role of AMPK activation in thyroid physiology and pathophysiology, with special focus on thyroid cancer.

High-energy compounds mobilize intracellular Ca2+ and activate calpain in cultured cells: Is calpain an energy-dependent protease?

Deficiency in energy metabolisms is perhaps the earliest modifiable defect in brain aging and sporadic Alzheimer's disease (sAD). Several high-energy compounds (HECs) such as ATP, phosphoenolpyruvate, phosphocreatine and acetyl coenzyme A have been shown to exhibit neuroprotective effects. To understand their mechanism of actions, we tested the effects of these HECs on intracellular Ca(2+), a central regulator in brain function. Our data showed that the HECs robustly and dose-dependently mobilized intracellular Ca(2+) in cultured SH-SY5Y cells, and the actions were sensitive to intracellular Ca(2+) chelator BAPTA-AM or energy metabolism blocker rotenone. The Ca(2+) influx triggered by the HECs was from both extracellular medium and intracellular stores and the HECs also induced repetitive Ca(2+) oscillations. As these actions were similar to those of classical Ca(2+) agonists, the HECs may be viewed as a new group of physiological Ca(2+) agonists. We also found that the HECs promoted the intracellular activity of calpain, a Ca(2+)-dependent protease, and the enzyme activity fluctuated in concert with cellular energy levels, suggesting that calpain activity may also be energy-driven or energy-dependent. These findings may add to current knowledge for the regulatory mechanisms of Ca(2+) and calpain. Since Ca(2+) and calpain undergo critical dysfunction in brain aging but the underlying mechanisms remain elusive, our work may provide a new perspective for clarifying some controversies. More importantly, the HECs, as key intermediates in glucose catabolism, the primary source of energy supply in the brain, may be used as potential drugs for rational prevention of sAD.

AMPK: a cellular energy sensor primarily regulated by AMP

AMPK (AMP-activated protein kinase) is a cellular energy sensor that monitors the ratio of AMP/ATP, and possibly also ADP/ATP, inside cells. Once activated by falling cellular energy levels, it acts to restore energy homoeostasis by switching on catabolic pathways that generate ATP, while switching off anabolic pathways and other processes consuming ATP. AMPK is switched on by increases in AMP via three mechanisms, all of which are antagonized by ATP: (i) promotion of phosphorylation of Thr172 by upstream activating kinases; (ii) inhibition of dephosphorylation of Thr172 by phosphatases; and (iii) allosteric activation of the phosphorylated kinase. Recently, it has been proposed that the first two mechanisms are also triggered by ADP, which might be the physiological signal rather than AMP, and that the third mechanism may not be physiologically significant. We have re-evaluated these questions, and found that only mechanism (ii) is mimicked by ADP, and that ADP is also less potent than AMP, which we still believe to be the primary signal. We have also provided evidence that mechanism (iii), i.e. allosteric activation by AMP, is a quantitatively significant mechanism in intact cells.

Targeting mTOR network in colorectal cancer therapy.

The mechanistic target of rapamycin (mTOR) integrates growth factor signals with cellular nutrient and energy levels and coordinates cell growth, proliferation and survival. A regulatory network with multiple feedback loops has evolved to ensure the exquisite regulation of cell growth and division. Colorectal cancer is the most intensively studied cancer because of its high incidence and mortality rate. Multiple genetic alterations are involved in colorectal carcinogenesis, including oncogenic Ras activation, phosphatidylinositol 3-kinase pathway hyperactivation, p53 mutation, and dysregulation of wnt pathway. Many oncogenic pathways activate the mTOR pathway. mTOR has emerged as an effective target for colorectal cancer therapy. In vitro and preclinical studies targeting the mTOR pathway for colorectal cancer chemotherapy have provided promising perspectives. However, the overall objective response rates in major solid tumors achieved with single-agent rapalog therapy have been modest, especially in advanced metastatic colorectal cancer. Combination regimens of mTOR inhibitor with agents such as cytotoxic chemotherapy, inhibitors of vascular endothelial growth factor, epidermal growth factor receptor and Mitogen-activated protein kinase kinase (MEK) inhibitors are being intensively studied and appear to be promising. Further understanding of the molecular mechanism in mTOR signaling network is needed to develop optimized therapeutic regimens. In this paper, oncogenic gene alterations in colorectal cancer, as well as their interaction with the mTOR pathway, are systematically summarized. The most recent preclinical and clinical anticancer therapeutic endeavors are reviewed. New players in mTOR signaling pathway, such as non-steroidal anti-inflammatory drug and metformin with therapeutic potentials are also discussed here.

MTOR signaling in autophagy regulation in the kidney.

Cells possess adaptive biosynthetic systems to maintain cellular energy levels for survival under adverse environmental conditions. Autophagy is an evolutionarily conserved cellular catabolic process that breaks down and recycles cytosolic material including macromolecules and organelles through lysosomal degradation. This catabolic process, represented by macroautophagy, is induced by a variety of cellular stresses such as nutrient starvation, which causes a shortage of cellular energy for cells to maintain cellular homeostasis and essential biological activities. In contrast, upon nutrient availability, cells stimulate anabolic processes. The mechanistic/mammalian target of rapamycin, a serine/threonine protein kinase, is a key player in stimulating cellular anabolism in response to nutrients and growth factors, and plays a crucial role in suppressing autophagy activity. Growing evidence has suggested that autophagy activity is required for the maintenance and physiological functions of renal cells including proximal tubular cells and podocytes. In this article, we discuss recent progress in the regulation of autophagy by mechanistic/mammalian target of rapamycin signaling.

CD28 Regulates Metabolism In Multiple Myeloma For Cell Survival and Proliferation

Multiple myeloma is an incurable hematological malignancy of transformed plasma cells. Many cellular interactions and soluble factors have been demonstrated to play a role in myeloma pathogenesis; however, novel targets to enhance therapeutic intervention are needed. We have demonstrated that CD28 signaling in myeloma cells supports their survival during chemotherapeutic challenge in vitro and in vivo. However, the cellular mechanisms by which CD28 confers this survival advantage to myeloma cells are not completely understood.CD28 is best characterized as the canonical T cell co-stimulatory molecule. During T cell activation, CD28 signaling induces glycolysis, a metabolic program required for T cell proliferation and functional maturation. In the absence of glycolysis, T cells utilize fatty acid oxidation for energy production through the mitochondria. However, the way in which CD28 regulates metabolism in multiple myeloma is not well understood. Here we present evidence that CD28 signaling induces glut1 expression, and that poisoning the glycolytic pathway inhibits proliferation and survival of myeloma cells.AMPK, an energy sensitive kinase known to regulate metabolism by driving fatty acid oxidation, is normally activated when cellular energy levels are low. Interestingly, poisoning glycolysis with a glucose analogue that cannot be processed (2DG) leads to AMPK inhibition in myeloma cells. Furthermore, pharmacological activation of AMPK by AICAR, an AMP analogue, is not sufficient to rescue myeloma cell proliferation from glycolytic inhibition and in fact increases cell death (p<.01 from no treatment, p<.05 from 2DG). This evidence suggests that multiple myeloma cells are absolutely dependent upon CD28-mediated glycolysis for proliferation and survival, and that myeloma cells cannot utilize fatty acid oxidation as a subsidiary metabolic pathway for proliferation in the absence of glycolysis. This understanding will allow us to target metabolism in multiple myeloma as a novel therapeutic strategy through pharmacological targeting of the CD28 pathway. This approach can be quickly translated into the clinic, as there are FDA approved drugs which activate AMPK (Metformin) and block CD28 signaling (Abatacept). Disclosures:Off Label Use: Abatacept, purpose to prevent CD28-mediated cell survival in multiple myeloma Metformin, purpose to activate AMPK in driving multiple myeloma cell death.

Regulation of SIRT1 by MicroRNAs

Sirtuin 1 (SIRT1) is an NAD(+)-dependent deacetylase that connects cellular energy levels to homeostatic responses by deacetylating and modulating the activities of many transcriptional regulators. Discovered as a longevity protein in yeast, the mammalian SIRT1 has been intensively studied because of its great potential as a therapeutic target to benefit human health by preventing and improving many age-related diseases. There has been, therefore, substantial interest in developing agents that upregulate SIRT1 expression and activity. SIRT1 is regulated at multiple levels, including post-transcriptionally by microRNAs (miRs), powerful regulators of diverse biological pathways. Here we discuss how expression and activity of SIRT1 and other sirtuins are inhibited by miRs and further discuss the therapeutic potential of targeting miRs for age-related diseases that involve SIRT1 dysfunction, focusing on obesityrelated diseases.

The systems biology of mitochondrial fission and fusion and implications for disease and aging

Mitochondria organize themselves as dynamic populations within a cell, by undergoing continuous cycles of fission and fusion. The spatio-temporal distribution and abundance of mitochondria determines the cell’s energy budget and is thus intimately linked to the cell’s response to environmental stimuli during aging. The dynamic balance of mitochondrial fission and fusion can be studied in terms of antagonistic subpopulations that regulate the mitochondrial responses in space and time. The dynamic nature of these processes motivates mathematical modelling and the simulation of such complex process. In several neurodegenerative and metabolic diseases the dynamic balance of fission and fusion is disturbed. However, how this dynamics plays a role in the progression of diseases is largely unclear. Fission and fusion help mitochondria to regulate cellular energy (ATP) levels, and minimize accumulation of harmful oxidized material called reactive oxygen species which accelerate mutations in mitochondrial DNA (mtDNA) during aging. We discuss how systems biology approaches can be used to investigate the mechanisms controlling the fission–fusion dynamics under two categories: dissecting the design of its molecular regulatory motifs, and understanding complex mitochondrial responses through their population level interactions. This will help us to understand how different regulatory mechanisms regulate the ATP and mutation (mtDNA) landscape of mitochondria to a variety of environmental stimuli in order to maintain their function during aging.