scholarly journals An upstream open reading frame represses translation of chicken PPARγ transcript variant 1

2019 ◽  
Author(s):  
Yankai Chu ◽  
Jiaxin Huang ◽  
Guangwei Ma ◽  
Tingting Cui ◽  
Xiaohong Yan ◽  
...  
Keyword(s):  
Mrna Stability ◽  
Posttranscriptional Regulation ◽  
Promoter Activity ◽  
Underlying Mechanism ◽  
Open Reading Frame ◽  
Upstream Open Reading Frame ◽  
Peroxisome Proliferator ◽  
Peroxisome Proliferator Activated Receptor ◽  
Reading Frame

AbstractPeroxisome proliferator-activated receptor γ (PPARγ) is a master regulator of adipogenesis. The PPARγ gene produces various transcripts with different 5′-untranslated regions (5′ UTRs) because of alternative promoter usage and splicing. The 5′ UTR plays important roles in posttranscriptional gene regulation. However, to date, the regulatory role and underlying mechanism of 5′ UTRs in the posttranscriptional regulation of PPARγ expression remain largely unclear. In this study, we investigated the effects of 5′ UTRs on posttranscriptional regulation using reporter assays. Our results showed that the five PPARγ 5′ UTRs exerted different effects on reporter gene activity. Bioinformatics analysis showed that chicken PPARγ transcript 1 (PPARγ1) possessed an upstream open reading frame (uORF) in its 5′ UTR. Mutation analysis showed that a mutation in the uORF led to increased Renilla luciferase activity and PPARγ protein expression, but decreased Renilla luciferase and PPARγ1 mRNA expression. mRNA stability analysis using real-time RT-PCR showed that the uORF mutation did not interfere with mRNA stability, but promoter activity analysis of the cloned 5′ UTR showed that the uORF mutation reduced promoter activity. Furthermore, in vitro transcription/translation assays demonstrated that the uORF mutation markedly increased the translation of PPARγ1 mRNA. Collectively, our results indicate that the uORF represses the translation of chicken PPARγ1 mRNA.

scholarly journals Bitter Orange (Citrus aurantium Linné) Improves Obesity by Regulating Adipogenesis and Thermogenesis through AMPK Activation

Nutrients ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 1988 ◽  
Author(s):  
Park ◽  
Kim ◽  
Jung ◽  
Ahn ◽  
Kwak ◽  
...  
Keyword(s):  
Uncoupling Protein ◽  
Underlying Mechanism ◽  
Peroxisome Proliferator ◽  
Citrus Aurantium ◽  
Brown Adipocytes ◽  
Peroxisome Proliferator Activated Receptor ◽  
Bitter Orange ◽  
Enhancer Binding Protein ◽  
Amp Activated Protein Kinase

Obesity is a global health threat. Herein, we evaluated the underlying mechanism of anti-obese features of bitter orange (Citrus aurantium Linné, CA). Eight-week-administration of CA in high fat diet-induced obese C57BL/6 mice resulted in a significant decrease of body weight, adipose tissue weight and serum cholesterol. In further in vitro studies, we observed decreased lipid droplets in CA-treated 3T3-L1 adipocytes. Suppressed peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer binding protein alpha indicated CA-inhibited adipogenesis. Moreover, CA-treated primary cultured brown adipocytes displayed increased differentiation associated with elevation of thermogenic factors including uncoupling protein 1 and PPARγ coactivator 1 alpha as well. The effects of CA in both adipocytes were abolished in AMP-activated protein kinase alpha (AMPKα)-suppressed environments, suggesting the anti-adipogenic and pro-thermogenic actions of CA were dependent on AMPKα pathway. In conclusion, our results suggest CA as a potential anti-obese agent which regulates adipogenesis and thermogenesis via AMPKα.

Posttranscriptional Regulation by the Upstream Open Reading Frame of the PhosphoethanolamineN-Methyltransferase Gene

2006 ◽  
Vol 70 (9) ◽  
pp. 2330-2334 ◽  
Author(s):  
Tomoki TABUCHI ◽  
Tomoyuki OKADA ◽  
Tetsushi AZUMA ◽  
Takashi NANMORI ◽  
Takeshi YASUDA
Keyword(s):  
Posttranscriptional Regulation ◽  
Open Reading Frame ◽  
Upstream Open Reading Frame ◽  
Reading Frame ◽  
Methyltransferase Gene

scholarly journals Effect of chronic contractile activity on mRNA stability in skeletal muscle

2010 ◽  
Vol 299 (1) ◽  
pp. C155-C163 ◽  
Author(s):  
Ruanne Y. J. Lai ◽  
Vladimir Ljubicic ◽  
Donna D'souza ◽  
David A. Hood
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Biogenesis ◽  
Mrna Stability ◽  
Degradation Kinetics ◽  
Contractile Activity ◽  
Peroxisome Proliferator ◽  
Turnover Rates ◽  
Peroxisome Proliferator Activated Receptor ◽  
Free System

Repeated bouts of exercise promote the biogenesis of mitochondria by multiple steps in the gene expression patterning. The role of mRNA stability in controlling the expression of mitochondrial proteins is relatively unexplored. To induce mitochondrial biogenesis, we chronically stimulated (10 Hz; 3 or 6 h/day) rat muscle for 7 days. Chronic contractile activity (CCA) increased the protein expression of PGC-1α, c-myc, and mitochondrial transcription factor A (Tfam) by 1.6-, 1.7- and 2.0-fold, respectively. To determine mRNA stability, we incubated total RNA with cytosolic extracts using an in vitro cell-free system. We found that the intrinsic mRNA half-lives ( t1/2) were variable within control muscle. Peroxisome proliferator-activated receptor-γ, coactivator-1α (PGC-1α) and Tfam mRNAs decayed more rapidly ( t1/2 = 22.7 and 31.4 min) than c-myc mRNA ( t1/2 = 99.7 min). Furthermore, CCA resulted in a differential response in degradation kinetics. After CCA, PGC-1α and Tfam mRNA half-lives decreased by 48% and 44%, respectively, whereas c-myc mRNA half-life was unchanged. CCA induced an elevation of both the cytosolic RNA-stabilizing human antigen R (HuR) and destabilizing AUF1 (total) by 2.4- and 1.8-fold, respectively. Increases in the p37AUF1, p40AUF1, and p45AUF1 isoforms were most evident. Thus these data indicate that CCA results in accelerated turnover rates of mRNAs encoding important mitochondrial biogenesis regulators in skeletal muscle. This adaptation is likely beneficial in permitting more rapid phenotypic plasticity in response to subsequent contractile activity.

scholarly journals Immunoregulation of Microglia M2 polarization is an unrecognized physiological function of α -Synuclein

2020 ◽  
Author(s):  
Na Li ◽  
Tessandra Stewart ◽  
Lifu Sheng ◽  
Min Shi ◽  
Eugene M Cilento ◽  
...  
Keyword(s):  
Lewy Body ◽  
Physiological Function ◽  
Physiological State ◽  
Underlying Mechanism ◽  
Peroxisome Proliferator ◽  
Peroxisome Proliferator Activated Receptor ◽  
Cellular Environment ◽  
Anti Inflammatory

Abstract BACKGROUNDMicroglial function is vital for maintaining the health of the brain, and their activation is an essential component of neurodegeneration. It is increasingly recognized that microglia also undergo changes, dependent on the cellular environment, that promote mainly reconstructive and anti-inflammatory functions, i.e. mostly desirable functions of microglia in a physiological state. What maintains microglia at this physiological state is essentially unknown, despite significant research on factors that provoke “reactive” or “inflammatory” phenotypes in conditions of injury or disease. One such factor, exposure to the aggregated or oligomeric forms of α-synuclein, an abundant brain protein, plays an essential role in driving microglial activation; including chemotactic migration and production of inflammatory mediators in Lewy body (LB) diseases such as Parkinson’s disease.METHODSIn this study, using in vitro and in vivo models, we challenged primary microglia or BV2 microglia with LPS + IFN-γ,IL-4 + IL-13, α-synuclein monomer and α-synuclein oligomer, examined microglia phenotype and the underlying mechanism by RT-PCR, Western blot, ELISA, IF, IHC, Co-IP. RESULTS: We described a novel physiological function of α-synuclein, in which it modulates microglia towards an anti-inflammatory phenotype by interaction with extracellular signal-regulated kinase (ERK) and recruitment of the ERK, nuclear factor kappa B (NF-κB), and peroxisome proliferator-activated receptor γ (PPARγ) pathways. CONCLUSIONS: These findings suggest a previously unrecognized function of α-synuclein that likely gives new insights into the pathogenesis and potential therapies for Lewy body-related diseases and beyond, given the abundance and multiple functions of α-synuclein in brain tissue.

scholarly journals microRNA-27a-3p but Not -5p Is a Crucial Mediator of Human Adipogenesis

Cells ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 3205
Author(s):  
Hang Wu ◽  
Taner Pula ◽  
Daniel Tews ◽  
Ez-Zoubir Amri ◽  
Klaus-Michael Debatin ◽  
...  
Keyword(s):  
Posttranscriptional Regulation ◽  
Adipogenic Differentiation ◽  
Regulation Of Gene Expression ◽  
Inhibitory Effect ◽  
Peroxisome Proliferator ◽  
Peroxisome Proliferator Activated Receptor ◽  
Inflammatory Conditions ◽  
Gene Assay ◽  
Sgbs Cells

MicroRNAs (miRNAs), a class of small, non-coding RNA molecules, play an important role in the posttranscriptional regulation of gene expression, thereby influencing important cellular functions. In adipocytes, miRNAs show import regulatory features and are described to influence differentiation as well as metabolic, endocrine, and inflammatory functions. We previously identified miR-27a being upregulated under inflammatory conditions in human adipocytes and aimed to elucidate its function in adipocyte biology. Both strands of miR-27a, miR-27a-3p and -5p, were downregulated during the adipogenic differentiation of Simpson–Golabi–Behmel syndrome (SGBS) cells, human multipotent adipose-derived stem cells (hMADS), and human primary adipose-derived stromal cells (hASCs). Using miRNA-mimic transfection, we observed that miR-27a-3p is a crucial regulator of adipogenesis, while miR-27a-5p did not alter the differentiation capacity in SGBS cells. In silico screening predicted lipoprotein lipase (LPL) and peroxisome proliferator activated receptor γ (PPARγ) as potential targets of miR-27a-3p. The downregulation of both genes was verified in vitro, and the interaction of miR-27-3p with target sites in the 3′ UTRs of both genes was confirmed via a miRNA-reporter-gene assay. Here, the knockdown of LPL did not interfere with adipogenic differentiation, while PPARγ knockdown decreased adipogenesis significantly, suggesting that miR-27-3p exerts its inhibitory effect on adipogenesis by repressing PPARγ. Taken together, we identified and validated a crucial role for miR-27a-3p in human adipogenesis played by targeting the essential adipogenic transcription factor PPARγ. Though we confirmed LPL as an additional target of miR-27a-3p, it does not appear to be involved in regulating human adipogenesis. Thereby, our findings call the conclusions drawn from previous studies, which identified LPL as a crucial regulator for murine and human adipogenesis, into question.

scholarly journals KLF2 Inhibits Chicken Preadipocyte Differentiation at Least in Part via Directly Repressing PPARγ Transcript Variant 1 Expression

2021 ◽  
Vol 9 ◽  
Author(s):  
Tingting Cui ◽  
Jiaxin Huang ◽  
Yingning Sun ◽  
Bolin Ning ◽  
Fang Mu ◽  
...  
Keyword(s):  
Binding Site ◽  
Promoter Activity ◽  
Methylation Status ◽  
Underlying Mechanism ◽  
Protein Isoforms ◽  
Transcript Variant ◽  
Peroxisome Proliferator ◽  
Preadipocyte Differentiation ◽  
Peroxisome Proliferator Activated Receptor ◽  
Endogenous Expression

Peroxisome proliferator-activated receptor gamma (PPARγ) is the master regulatory factor of preadipocyte differentiation. As a result of alternative splicing and alternative promoter usage, PPARγ gene generates multiple transcript variants encoding two protein isoforms. Krüppel-like factor 2 (KLF2) plays a negative role in preadipocyte differentiation. However, its underlying mechanism remains incompletely understood. Here, we demonstrated that KLF2 inhibited the P1 promoter activity of the chicken PPARγ gene. Bioinformatics analysis showed that the P1 promoter harbored a conserved putative KLF2 binding site, and mutation analysis showed that the KLF2 binding site was required for the KLF2-mediated transcription inhibition of the P1 promoter. ChIP, EMSA, and reporter gene assays showed that KLF2 could directly bind to the P1 promoter regardless of methylation status and reduced the P1 promoter activity. Consistently, histone modification analysis showed that H3K9me2 was enriched and H3K27ac was depleted in the P1 promoter upon KLF2 overexpression in ICP1 cells. Furthermore, gene expression analysis showed that KLF2 overexpression reduced the endogenous expression of PPARγ transcript variant 1 (PPARγ1), which is driven by the P1 promoter, in DF1 and ICP1 cells, and that the inhibition of ICP1 cell differentiation by KLF2 overexpression was accompanied by the downregulation of PPARγ1 expression. Taken together, our results demonstrated that KLF2 inhibits chicken preadipocyte differentiation at least inpart via direct downregulation of PPARγ1 expression.

scholarly journals Termination and Peptide Release at the Upstream Open Reading Frame Are Required for Downstream Translation on Synthetic Shunt-Competent mRNA Leaders

2000 ◽  
Vol 20 (17) ◽  
pp. 6212-6223 ◽  
Author(s):  
Maja Hemmings-Mieszczak ◽  
Thomas Hohn ◽  
Thomas Preiss
Keyword(s):  
Stop Codon ◽  
Cauliflower Mosaic Virus ◽  
Open Reading Frame ◽  
Upstream Open Reading Frame ◽  
Reading Frame ◽  
Codon Context ◽  
Necessary And Sufficient ◽  
Synthetic Mrna

ABSTRACT We have shown recently that a stable hairpin preceded by a short upstream open reading frame (uORF) promotes nonlinear ribosome migration or ribosome shunt on a synthetic mRNA leader (M. Hemmings-Mieszczak and T. Hohn, RNA 5:1149–1157, 1999). We have now used the model mRNA leader to study further the mechanism of shunting in vivo and in vitro. We show that a full cycle of translation of the uORF, including initiation, elongation, and termination, is a precondition for the ribosome shunt across the stem structure to initiate translation downstream. Specifically, AUG recognition and the proper release of the nascent peptide are necessary and sufficient for shunting. Furthermore, the stop codon context must not impede downstream reinitiation. Translation of the main ORF was inhibited by replacement of the uORF by coding sequences repressing reinitiation but stimulated by the presence of the virus-specific translational transactivator of reinitiation (cauliflower mosaic virus pVI). Our results indicate reinitiation as the mechanism of translation initiation on the synthetic shunt-competent mRNA leader and suggest that uORF-dependent shunting is more prevalent than previously anticipated. Within the above constraints, uORF-dependent shunting is quite tolerant of uORF and stem sequences and operates in systems as diverse as plants and fungi.

scholarly journals Smart-ORF: a single-molecule method for accessing ribosome dynamics in both upstream and main open reading frames

2020 ◽  
Author(s):  
Anthony Gaba ◽  
Hongyun Wang ◽  
Trinisia Fortune ◽  
Xiaohui Qu
Keyword(s):  
Single Molecule ◽  
Translational Regulation ◽  
Mrna Translation ◽  
Open Reading Frames ◽  
Open Reading Frame ◽  
Upstream Open Reading Frame ◽  
Translation Efficiency ◽  
Reading Frame ◽  
Reading Frames

Abstract Upstream open reading frame (uORF) translation disrupts scanning 43S flux on mRNA and modulates main open reading frame (mORF) translation efficiency. Current tools, however, have limited access to ribosome dynamics in both upstream and main ORFs of an mRNA. Here, we develop a new two-color in vitro fluorescence assay, Smart-ORF, that monitors individual uORF and mORF translation events in real-time with single-molecule resolution. We demonstrate the utility of Smart-ORF by applying it to uORF-encoded arginine attenuator peptide (AAP)-mediated translational regulation. The method enabled quantification of uORF and mORF initiation efficiencies, 80S dwell time, polysome formation, and the correlation between uORF and mORF translation dynamics. Smart-ORF revealed that AAP-mediated 80S stalling in the uORF stimulates the uORF initiation efficiency and promotes clustering of slower uORF-translating ribosomes. This technology provides a new tool that can reveal previously uncharacterized dynamics of uORF-containing mRNA translation.

scholarly journals Adipocyte PHLPP2 inhibition prevents obesity-induced fatty liver

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
KyeongJin Kim ◽  
Jin Ku Kang ◽  
Young Hoon Jung ◽  
Sang Bae Lee ◽  
Raffaela Rametta ◽  
...  
Keyword(s):  
Fatty Liver ◽  
Whole Body ◽  
Acid Oxidation ◽  
Chow Diet ◽  
Peroxisome Proliferator ◽  
Obese Mice ◽  
Ph Domain ◽  
Peroxisome Proliferator Activated Receptor

AbstractIncreased adiposity confers risk for systemic insulin resistance and type 2 diabetes (T2D), but mechanisms underlying this pathogenic inter-organ crosstalk are incompletely understood. We find PHLPP2 (PH domain and leucine rich repeat protein phosphatase 2), recently identified as the Akt Ser473 phosphatase, to be increased in adipocytes from obese mice. To identify the functional consequence of increased adipocyte PHLPP2 in obese mice, we generated adipocyte-specific PHLPP2 knockout (A-PHLPP2) mice. A-PHLPP2 mice show normal adiposity and glucose metabolism when fed a normal chow diet, but reduced adiposity and improved whole-body glucose tolerance as compared to Cre- controls with high-fat diet (HFD) feeding. Notably, HFD-fed A-PHLPP2 mice show increased HSL phosphorylation, leading to increased lipolysis in vitro and in vivo. Mobilized adipocyte fatty acids are oxidized, leading to increased peroxisome proliferator-activated receptor alpha (PPARα)-dependent adiponectin secretion, which in turn increases hepatic fatty acid oxidation to ameliorate obesity-induced fatty liver. Consistently, adipose PHLPP2 expression is negatively correlated with serum adiponectin levels in obese humans. Overall, these data implicate an adipocyte PHLPP2-HSL-PPARα signaling axis to regulate systemic glucose and lipid homeostasis, and suggest that excess adipocyte PHLPP2 explains decreased adiponectin secretion and downstream metabolic consequence in obesity.

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