Homeostasis

Summary

In biology, homeostasis (British also homoeostasis) (/hɒmɪə(ʊ)ˈsteɪsɪs/) is the state of steady internal, physical, chemical, and social conditions maintained by living systems. This is the condition of optimal functioning for the organism and includes many variables, such as body temperature and fluid balance, being kept within certain pre-set limits (homeostatic range). Other variables include the pH of extracellular fluid, the concentrations of sodium, potassium, and calcium ions, as well as the blood sugar level, and these need to be regulated despite changes in the environment, diet, or level of activity. Each of these variables is controlled by one or more regulators or homeostatic mechanisms, which together maintain life. Homeostasis is brought about by a natural resistance to change when already in optimal conditions, and equilibrium is maintained by many regulatory mechanisms; it is thought to be the central motivation for all organic action. All homeostatic control mechanisms have at least three interdependent components for the variable being regulated: a receptor, a control center, and an effector. The receptor is the sensing component that monitors and responds to changes in the environment, either external or internal. Receptors include thermoreceptors and mechanoreceptors. Control centers include the respiratory center and the renin-angiotensin system. An effector is the target acted on, to bring about the change back to the normal state. At the cellular level, effectors include nuclear receptors that bring about changes in gene expression through up-regulation or down-regulation and act in negative feedback mechanisms. An example of this is in the control of bile acids in the liver. Some centers, such as the renin–angiotensin system, control more than one variable. When the receptor senses a stimulus, it reacts by sending action potentials to a control center. The control center sets the maintenance range—the acceptable upper and lower limits—for the particular variable, such as temperature.

Mechanistic insights into AMP-activated protein kinase-dependent gene expression

Caterina Collodet

AMP-activated protein kinase (AMPK) is a fundamental enzyme that controls energy homeostasis, through orchestrating the cellular response to a reduction in energy availability. Under conditions of cellular energy stress AMPK senses the decrease in ATP levels and responds by activating catabolic pathways, which will generate ATP, and switching off ATP-consuming ones, in order to restore the energy balance. AMPK regulates several signaling cascades linked to metabolism, overall favoring cellular consumption of glucose and lipid, allowing the cell to adapt to sustained energetic challenges through modulation of gene transcription. The aim of this thesis is to investigate the role that AMPK plays in the adaptive reprogramming of metabolism through transcriptional control. To identify genes and pathways regulated in an AMPK-dependent mechanism, we performed a whole-genome transcriptome profiling using microarray technology and compared the effects of two small molecule AMPK activators acting via distinct mechanisms, namely 991, which binds at the allosteric drug and metabolite site, and the AMP mimetic, 5-aminoimidazole-4-carboxamide-1-Î²-D-ribofuranoside (AICAR). The impact on gene expression of 991 and AICAR was investigated using two cellular and genetic models, mouse embryonic fibroblasts (MEFs) and mouse primary hepatocytes, either wild-type or AMPK-deficient. Statistical analysis of differential gene expression, followed by pathway analysis, revealed compound- and model-specific gene expression signatures. Notably we found that in contrast to AICAR, 991 affected gene expression almost exclusively in an AMPK-dependent manner. Interestingly, we identified that 991 modulated genes involved in the metabolic and lysosomal pathways, and that a number of these genes are under the control of the sterol regulatory element-binding protein (SREBP) and transcription factor EB (TFEB). We identified the tumor suppressor folliculin (Flcn) and its binding partners, folliculin interacting protein (Fnip), as novel transcriptional targets of AMPK. We confirmed the upregulated expression of Flcn in response to pharmacological activation of AMPK in MEFs and primary hepatocytes. Furthermore, by taking advantage of a novel zebrafish whole-body knockout model of AMPK, we confirmed that physiological activation of AMPK partly mediates the increase in expression of Flcn and Fnip. We further identified TFEB as a mediator of the AMPK transcriptional response, accounting for the increase in Flcn expression through modulating its promoter activity. Moreover, we revealed the existence of a novel mechanism by which AMPK regulates TFEB through promoting dephosphorylation and nuclear translocation, independently of mammalian target of rapamycin complex 1 (mTORC1). Taken together, we identified several new AMPK-dependent/-regulated genes and pathways that are differentially modulated in a cell type- and compound-specific manner. Most importantly, we discovered a novel and conserved AMPK-TFEB-FLCN axis in cellular and in vivo models. This work contributes to advance our understanding of AMPK-mediated regulation of transcriptional programs, nevertheless future studies will be required to elucidate the physiological relevance of the AMPK-TFEB-FLCN cascade.

EPFL2019

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AMP-activated protein kinase (AMPK) plays diverse roles and coordinates complex metabolic pathways for maintenance of energy homeostasis. This could be explained by the fact that AMPK exists as multiple heterotrimer complexes comprising a catalytic alpha-subunit (alpha 1 and alpha 2) and regulatory beta (beta 1 and beta 2)- and gamma (gamma 1, gamma 2, gamma 3)-subunits, which are uniquely distributed across different cell types. There has been keen interest in developing specific and isoform-selective AMPK-activating drugs for therapeutic use and also as research tools. Moreover, establishing ways of enhancing cellular AMPK activity would be beneficial for both purposes. Here, we investigated if a recently described potent AMPK activator called 991, in combination with the commonly used activator 5-aminoimidazole-4-carboxamide riboside or contraction, further enhances AMPK activity and glucose transport in mouse skeletal muscle ex vivo. Given that the gamma 3-subunit is exclusively expressed in skeletal muscle and has been implicated in contraction-induced glucose transport, we measured the activity of AMPK gamma 3 as well as ubiquitously expressed gamma 1-containing complexes. We initially validated the specificity of the antibodies for the assessment of isoform-specific AMPK activity using AMPK-deficient mouse models. We observed that a low dose of 991 (5 mu M) stimulated a modest or negligible activity of both gamma 1- and gamma 3-containing AMPK complexes. Strikingly, dual treatment with 991 and 5-aminoimidazole-4-carboxamide riboside or 991 and contraction profoundly enhanced AMPK gamma 1/gamma 3 complex activation and glucose transport compared with any of the single treatments. The study demonstrates the utility of a dual activator approach to achieve a greater activation of AMPK and downstream physiological responses in various cell types, including skeletal muscle.

Amer Physiological Soc2016

Enhanced activation of cellular AMPK by dual-small molecule treatment: AICAR and A769662

Kei Sakamoto, Maria Deak, Laurent Bultot

AMP-activated protein kinase (AMPK) is a key cellular energy sensor and regulator of metabolic homeostasis. Activation of AMPK provides beneficial outcomes in fighting against metabolic disorders such as insulin resistance and type 2 diabetes. Currently, there is no allosteric AMPK activator available for the treatment of metabolic diseases, and limited compounds are available to robustly stimulate cellular/tissue AMPK in a specific manner. Here we investigated whether simultaneous administration of two different pharmacological AMPK activators, which bind and act on different sites, would result in an additive or synergistic effect on AMPK and its downstream signaling and physiological events in intact cells. We observed that cotreating primary hepatocytes with the AMP mimetic 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR) and a low dose (1 mu M) of the allosteric activator A769662 produced a synergistic effect on AMPK Thr172 phosphorylation and catalytic activity, which was associated with a more profound increase/decrease in phosphorylation of downstream AMPK targets and inhibition of hepatic lipogenesis compared with single-compound treatment. Mechanistically, we found that co-treatment does not stimulate LKB1, upstream kinase for AMPK, but it protects against dephosphorylation of Thr172 phosphorylation by protein phosphatase PP2C alpha in an additive manner in a cell-free assay. Collectively, we demonstrate that AICAR sensitizes the effect of A769662 and promotes AMPK activity and its downstream events. The study demonstrates the feasibility of promoting AMPK activity by using two activators with distinct modes of action in order to achieve a greater activation of AMPK and downstream signaling.

Amer Physiological Soc2014