scholarly journals LXRα Phosphorylation in Cardiometabolic Disease: Insight From Mouse Models

Endocrinology ◽  
2020 ◽  
Vol 161 (7) ◽  
Author(s):  
Maud Voisin ◽  
Matthew C Gage ◽  
Natalia Becares ◽  
Elina Shrestha ◽  
Edward A Fisher ◽  
...  
Keyword(s):  
Nuclear Receptors ◽  
Posttranslational Modifications ◽  
Physiological Response ◽  
Metabolic Diseases ◽  
Cardiometabolic Disease ◽  
Nonalcoholic Fatty Liver ◽  
Environmental Signals ◽  
Powerful Means ◽  
And Function

Abstract Posttranslational modifications, such as phosphorylation, are a powerful means by which the activity and function of nuclear receptors such as LXRα can be altered. However, despite the established importance of nuclear receptors in maintaining metabolic homeostasis, our understanding of how phosphorylation affects metabolic diseases is limited. The physiological consequences of LXRα phosphorylation have, until recently, been studied only in vitro or nonspecifically in animal models by pharmacologically or genetically altering the enzymes enhancing or inhibiting these modifications. Here we review recent reports on the physiological consequences of modifying LXRα phosphorylation at serine 196 (S196) in cardiometabolic disease, including nonalcoholic fatty liver disease, atherosclerosis, and obesity. A unifying theme from these studies is that LXRα S196 phosphorylation rewires the LXR-modulated transcriptome, which in turn alters physiological response to environmental signals, and that this is largely distinct from the LXR-ligand–dependent action.

scholarly journals Metabolic endotoxemia promotes adipose dysfunction and inflammation in human obesity

2019 ◽  
Vol 316 (2) ◽  
pp. E319-E332 ◽  
Author(s):  
Mercedes Clemente-Postigo ◽  
Wilfredo Oliva-Olivera ◽  
Leticia Coin-Aragüez ◽  
Bruno Ramos-Molina ◽  
Rosa María Giraldez-Perez ◽  
...  
Keyword(s):  
Metabolic Diseases ◽  
In Vitro Assays ◽  
Low Grade ◽  
Human Obesity ◽  
Gene And Protein Expression ◽  
And Function ◽  
Adipose Dysfunction ◽  
High Degree ◽  
Metabolic Endotoxemia

Impaired adipose tissue (AT) lipid handling and inflammation is associated with obesity-related metabolic diseases. Circulating lipopolysaccharides (LPSs) from gut microbiota (metabolic endotoxemia), proposed as a triggering factor for the low-grade inflammation in obesity, might also be responsible for AT dysfunction. Nevertheless, this hypothesis has not been explored in human obesity. To analyze the relationship between metabolic endotoxemia and AT markers for lipogenesis, lipid handling, and inflammation in human obesity, 33 patients with obesity scheduled for surgery were recruited and classified according to their LPS levels. Visceral and subcutaneous AT gene and protein expression were analyzed and adipocyte and AT in vitro assays performed. Subjects with obesity with a high degree of metabolic endotoxemia had lower expression of key genes for AT function and lipogenesis ( SREBP1, FABP4, FASN, and LEP) but higher expression of inflammatory genes in visceral and subcutaneous AT than subjects with low LPS levels. In vitro experiments corroborated that LPS are responsible for adipocyte and AT inflammation and downregulation of PPARG, SCD, FABP4, and LEP expression and LEP secretion. Thus, metabolic endotoxemia influences AT physiology in human obesity by decreasing the expression of factors involved in AT lipid handling and function as well as by increasing inflammation.

scholarly journals Posttranslational Modifications and the Immunogenicity of Biotherapeutics

2016 ◽  
Vol 2016 ◽  
pp. 1-15 ◽  
Author(s):  
Roy Jefferis
Keyword(s):  
Posttranslational Modifications ◽  
Drug Product ◽  
Molecular Forms ◽  
Phagocytic Cells ◽  
Production Platform ◽  
The Individual ◽  
And Function ◽  
Antigen Presenting

Whilst the amino acid sequence of a protein is determined by its gene sequence, the final structure and function are determined by posttranslational modifications (PTMs), including quality control (QC) in the endoplasmic reticulum (ER) and during passage through the Golgi apparatus. These processes are species and cell specific and challenge the biopharmaceutical industry when developing a production platform for the generation of recombinant biologic therapeutics. Proteins and glycoproteins are also subject to chemical modifications (CMs) bothin vivoandin vitro. The individual is naturally tolerant to molecular forms of self-molecules but nonself variants can provoke an immune response with the generation of anti-drug antibodies (ADA); aggregated forms can exhibit enhanced immunogenicity and QC procedures are developed to avoid or remove them. Monoclonal antibody therapeutics (mAbs) are a special case because their purpose is to bind the target, with the formation of immune complexes (ICs), a particular form of aggregate. Such ICs may be removed by phagocytic cells that have antigen presenting capacity. These considerations may frustrate the possibility of ameliorating the immunogenicity of mAbs by rigorous exclusion of aggregates from drug product. Alternate strategies for inducing immunosuppression or tolerance are discussed.

scholarly journals Phosphorylation of Steroidogenic Factor 1 Is Mediated by Cyclin-Dependent Kinase 7

2008 ◽  
Vol 22 (1) ◽  
pp. 91-104 ◽  
Author(s):  
Aurélia E. Lewis ◽  
Marte Rusten ◽  
Erling A. Hoivik ◽  
Elisabeth L. Vikse ◽  
Magnus L. Hansson ◽  
...  
Keyword(s):  
Posttranslational Modifications ◽  
Cdk Inhibitor ◽  
Structural Studies ◽  
Cyclin Dependent Kinase ◽  
Steroidogenic Factor 1 ◽  
Phosphorylation Level ◽  
Inactive Form ◽  
And Function ◽  
Cdk Activating Kinase

Abstract The nuclear receptor steroidogenic factor-1 (SF1) is critical for development and function of steroidogenic tissues. Posttranslational modifications are known to influence the transcriptional capacity of SF1, and it was previously demonstrated that serine 203 is phosphorylated. In this paper we report that serine 203 is phosphorylated by a cyclin-dependent kinase 7 (CDK7)-mediated process. As part of the CDK-activating kinase complex, CDK7 is a component of the basal transcription factor TFIIH, and phosphorylation of SF1 as well as SF1-dependent transcription was clearly reduced in cells carrying a mutation that renders the CDK-activating kinase complex unable to interact with the TFIIH core. Coimmunoprecipitation analyses revealed that SF1 and CDK7 reside in the same complex, and kinase assays demonstrated that immunoprecipitated CDK7 and purified TFIIH phosphorylate SF1 in vitro. The CDK inhibitor roscovitine blocked phosphorylation of SF1, and an inactive form of CDK7 repressed the phosphorylation level and the transactivation capacity of SF1. Structural studies have identified phosphoinositides as potential ligands for SF1. Interestingly, we found that mutations designed to block phospholipid binding dramatically decreased the level of SF1 phosphorylation. Together our results suggest a connection between ligand occupation and phosphorylation and association with the basic transcriptional machinery, indicating an intricate regulation of SF1 transactivation.

scholarly journals Site-specific phosphorylation of Huntingtin exon 1 recombinant proteins enabled by the discovery of novel kinases

2020 ◽  
Author(s):  
Anass Chiki ◽  
Jonathan Ricci ◽  
Ramanath Hegde ◽  
Luciano A. Abriata ◽  
Andreas Reif ◽  
...  
Keyword(s):  
Posttranslational Modifications ◽  
Proof Of Concept ◽  
Wild Type ◽  
Expression And Purification ◽  
Sumo Fusion ◽  
Exon 1 ◽  
Health And Disease ◽  
Experimental Approaches ◽  
And Function

AbstractPosttranslational modifications (PTMs) within the first 17 amino acids (Nt17) of exon1 of the Huntingtin protein (Httex1) play important roles in modulating its cellular properties and functions in health and disease. In particular, phosphorylation of threonine and serine residues (T3, S13, and/or S16) has been shown to inhibit Htt aggregation in vitro and inclusion formation in cellular and animal models of Huntington’s disease (HD). In this manuscript, we describe a new and simple methodology for producing milligram quantities of highly pure wild type or mutant Httex1 proteins that are site-specifically phosphorylated at T3 or at both S13 and S16. This advance was enabled by 1) the discovery and validation of novel kinases that efficiently phosphorylate Httex1 at S13 and S16 (TBK1), at T3 (GCK) or T3 and S13 (TNIK and HGK); and, 2) the development of an efficient methodology for producing recombinant native Httex1 proteins using a SUMO-fusion expression and purification strategy. As proof of concept, we demonstrate how this method can be applied to produce Httex1 proteins that are both site- specifically phosphorylated and fluorescently labeled or isotopically labeled. Together, these advances should increase access to these valuable tools and expand the range of methods and experimental approaches that can be used to elucidate the mechanisms by which phosphorylation influences Httex1 structure, aggregation, interactome and function(s) in health and disease.

scholarly journals Electrochemistry as a surrogate for protein phosphorylation: voltage-controlled assembly of reflectin A1

2020 ◽  
Vol 17 (173) ◽  
pp. 20200774
Author(s):  
Sheng-Ping Liang ◽  
Robert Levenson ◽  
Brandon Malady ◽  
Michael J. Gordon ◽  
Daniel E. Morse ◽  
...  
Keyword(s):  
Low Voltage ◽  
Selective Reduction ◽  
Structure And Function ◽  
Fine Tuning ◽  
Primary Amines ◽  
Environmental Signals ◽  
Instrument Measuring ◽  
And Function

Phosphorylation is among the most widely distributed mechanisms regulating the tunable structure and function of proteins in response to neuronal, hormonal and environmental signals. We demonstrate here that the low-voltage electrochemical reduction of histidine residues in reflectin A1, a protein that mediates the neuronal fine-tuning of colour reflected from skin cells for camouflage and communication in squids, acts as an in vitro surrogate for phosphorylation in vivo , driving the assembly previously shown to regulate its function. Using micro-drop voltammetry and a newly designed electrochemical cell integrated with an instrument measuring dynamic light scattering, we demonstrate selective reduction of the imidazolium side chains of histidine in monomers, oligopeptides and this complex protein in solution. The formal reduction potential of imidazolium proves readily distinguishable from those of hydronium and primary amines, allowing unequivocal confirmation of the direct and energetically selective deprotonation of histidine in the protein. The resulting ‘electro-assembly’ provides a new approach to probe, understand, and control the mechanisms that dynamically tune protein structure and function in normal physiology and disease. With its abilities to serve as a surrogate for phosphorylation and other mechanisms of charge neutralization, and to potentially isolate early intermediates in protein assembly, this method may be useful for analysing never-before-seen early intermediates in the phosphorylation-driven assembly of other proteins in normal physiology and disease.

scholarly journals Autophagy Induced by Palmitic Acid: a Brake in NAFLD Neutrophils

2021 ◽  
Author(s):  
Zhicheng Peng ◽  
Heyuan Wang ◽  
Alan Y. Hsu ◽  
Xiliang Du ◽  
Yuchen Yang ◽  
...  
Keyword(s):  
Palmitic Acid ◽  
Metabolic Diseases ◽  
Ex Vivo ◽  
Neutrophil Function ◽  
List Type ◽  
Nonalcoholic Fatty Liver ◽  
Blood Constituents ◽  
Key Points ◽  
Ethanol Toxicity

AbstractInnate immune suppression and high blood fatty acid levels are the pathological basis of multiple metabolic diseases. Neutrophil vacuolation is an indicator of the immune status of patients, which is associated with autophagy-dependent granule degradation. Vacuolated neutrophils are observed in ethanol toxicity and septicemia patients due to the changes in their blood constituents, but how about the neutrophils in nonalcoholic fatty liver disease (NAFLD) patient is unknown. Here, we confirmed that an adhesion deficiency and an increased autophagy level existed in NAFLD neutrophils, and the three neutrophil granule subunits, namely, the azurophil granules, specific granules and gelatinase granules, could be engulfed by autophagosomes for degradation, and these autophagy-triggered granule degradation events were associated with vacuolation in palmitic acid (PA)-treated and NAFLD neutrophils. Concordantly, the adhesion-associated molecules CD11a, CD11b, CD18 and Rap1 on the three granule subunits were degraded during PA induced autophagy. Moreover, the cytosolic CD11a, CD11b, CD18 and Rap1 were targeted by Hsc70 and then delivered to lysosomal-like granules for degradation. Notably, in vitro and ex vivo, PA induced autophagy by inhibiting the p-PKCα/PKD2 pathway. Overall, we showed that high blood PA level inhibited the p-PKCα/PKD2 pathway to induce NAFLD neutrophil autophagy, which promoted the degradation of CD11a, CD11b, CD18 and Rap1 and further decreased the adhesion of neutrophils, thereby impairing the neutrophil function of NAFLD patients. This theory provides a new therapeutic strategy to improve the immune deficiency in NAFLD patients.Visual AbstractKey PointsVacuolation and adhesion deficiency of NAFLD neutrophils are associated with autophagy-dependent granule degradationPA inhibits p-PKCα/PKD2 to induce autophagy, which induces the degradation of CD11a, CD11b, CD18 and Rap1 and decreases neutrophil adhesion

scholarly journals The interface of nuclear and membrane steroid signaling

Endocrinology ◽  
2021 ◽  
Author(s):  
Lindsey S Treviño ◽  
Daniel A Gorelick
Keyword(s):  
Estrogen Receptor ◽  
Transcription Factors ◽  
Membrane Proteins ◽  
Cell Membrane ◽  
Nuclear Receptors ◽  
Cell Nucleus ◽  
Androgen Receptors ◽  
Steroid Receptors ◽  
And Function

Abstract Steroid hormones bind receptors in the cell nucleus and in the cell membrane. The most widely studied class of steroid hormone receptors are the nuclear receptors, named for their function as ligand-dependent transcription factors in the cell nucleus. Nuclear receptors, such as estrogen receptor alpha, can also be anchored to the plasma membrane, where they respond to steroids by activating signaling pathways independent of their function as transcription factors. Steroids can also bind integral membrane proteins, such as the G protein-coupled estrogen receptor. Membrane estrogen and progestin receptors have been cloned and characterized in vitro and influence the development and function of many organ systems. Membrane androgen receptors were cloned and characterized in vitro, but their function as androgen receptors in vivo is unresolved. We review the identity and function of membrane proteins that bind estrogens, progestins and androgens. We discuss evidence that membrane glucocorticoid and mineralocorticoid receptors exist, and whether glucocorticoid and mineralocorticoid nuclear receptors act at the cell membrane. In many cases, integral membrane steroid receptors act independently of nuclear steroid receptors, even though they may share a ligand.

scholarly journals DOT1L Regulates Thermogenic Adipocyte Differentiation and Function via Modulating H3K79 Methylation

2021 ◽  
Author(s):  
Lin Shuai ◽  
Bo-Han Li ◽  
Hao-Wen Jiang ◽  
Lin Yang ◽  
Jia Li ◽  
...  
Keyword(s):  
Energy Homeostasis ◽  
Adipocyte Differentiation ◽  
Metabolic Diseases ◽  
Diet Induced Obesity ◽  
Beige Adipocytes ◽  
Thermogenic Adipocytes ◽  
And Function ◽  
H3k79 Methylation

Brown and beige adipocytes are characterized as thermogenic adipocytes and have great potential for treating obesity and associated metabolic diseases. Here, we identify a conserved mammalian lysine 79 of histone H3 (H3K79) methyltransferase, disruptor of telomeric silencing -1 like (DOT1L), as a new epigenetic regulator that controls thermogenic adipocyte differentiation and function. We show that deletion of DOT1L in thermogenic adipocytes potently protects mice from diet-induced obesity, improves glucose homeostasis, alleviates hepatic steatosis, and facilitates adaptive thermogenesis<i> in vivo</i>. Loss of DOT1L in primary preadipocytes significantly promotes brown and beige adipogenesis and thermogenesis<i> in vitro</i>. Mechanistically, DOT1L epigenetically regulates the BAT-selective gene program through modulating H3K79 methylation, in particular H3K79me2 modification. Thus, our study demonstrates that DOT1L exerts an important role in energy homeostasis by regulating thermogenic adipocyte differentiation and function.

scholarly journals Silencing of functional p53 attenuates NAFLD by promoting HMGB1-related autophagy induction

2020 ◽  
Vol 14 (5) ◽  
pp. 828-841
Author(s):  
Xuequn Zhang ◽  
Yiming Lin ◽  
Sisi Lin ◽  
Chunxiao Li ◽  
Jianguo Gao ◽  
...  
Keyword(s):  
Liver Disease ◽  
Metabolic Diseases ◽  
High Mobility ◽  
Cell Models ◽  
Tumor Suppressor P53 ◽  
Nonalcoholic Fatty Liver ◽  
Autophagy Induction ◽  
Huh7 Cells

Abstract Background and aim Nonalcoholic fatty liver disease (NAFLD) is a common chronic liver disease worldwide, but its pathogenesis remains imprecisely understood and requires further clarification. Recently, the tumor suppressor p53 has received growing attention for its role in metabolic diseases. In this study, we performed in vivo and in vitro experiments to identify the contribution of p53–autophagy regulation to NAFLD. Methods Livers from wild-type and p53 knockout mice as well as p53-functional HepG2 cells and p53-dysfunctional Huh7 cells were examined for autophagy status and HMGB1 translocation. In vivo and in vitro NAFLD models were established, and steatosis was detected. In the cell models, autophagy status and steatosis were examined by p53 and/or HMGB1 silencing. Results First, the silencing of p53 could induce autophagy both in vivo and in vitro. In addition, p53 knockout attenuated high-fat diet-induced NAFLD in mice. Similarly, knockdown of p53 could alleviate palmitate-induced lipid accumulation in cell models. Furthermore, high mobility group box 1 (HMGB1) was proven to contribute to the effect of silencing p53 on alleviating NAFLD in vitro as an autophagy regulator. Conclusion The anti-NAFLD effect of functional p53 silencing is associated with the HMGB1-mediated induction of autophagy. Graphic abstract

scholarly journals DOT1L Regulates Thermogenic Adipocyte Differentiation and Function via Modulating H3K79 Methylation

2021 ◽  
Author(s):  
Lin Shuai ◽  
Bo-Han Li ◽  
Hao-Wen Jiang ◽  
Lin Yang ◽  
Jia Li ◽  
...  
Keyword(s):  
Energy Homeostasis ◽  
Adipocyte Differentiation ◽  
Metabolic Diseases ◽  
Diet Induced Obesity ◽  
Beige Adipocytes ◽  
Thermogenic Adipocytes ◽  
And Function ◽  
H3k79 Methylation

Brown and beige adipocytes are characterized as thermogenic adipocytes and have great potential for treating obesity and associated metabolic diseases. Here, we identify a conserved mammalian lysine 79 of histone H3 (H3K79) methyltransferase, disruptor of telomeric silencing -1 like (DOT1L), as a new epigenetic regulator that controls thermogenic adipocyte differentiation and function. We show that deletion of DOT1L in thermogenic adipocytes potently protects mice from diet-induced obesity, improves glucose homeostasis, alleviates hepatic steatosis, and facilitates adaptive thermogenesis<i> in vivo</i>. Loss of DOT1L in primary preadipocytes significantly promotes brown and beige adipogenesis and thermogenesis<i> in vitro</i>. Mechanistically, DOT1L epigenetically regulates the BAT-selective gene program through modulating H3K79 methylation, in particular H3K79me2 modification. Thus, our study demonstrates that DOT1L exerts an important role in energy homeostasis by regulating thermogenic adipocyte differentiation and function.

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