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MuRF2 regulates PPARγ1 activity to protect against diabetic cardiomyopathy and enhance weight gain induced by a high fat diet.

He J, Quintana MT, Sullivan J, L Parry T, J Grevengoed T, Schisler JC, Hill JA, Yates CC, Mapanga RF, Essop MF, Stansfield WE, Bain JR, Newgard CB, Muehlbauer MJ, Han Y, Clarke BA, Willis MS - Cardiovasc Diabetol (2015)

Bottom Line: However, increasing MuRF2:PPARγ1 (>5:1) beyond physiological levels drove poly-ubiquitin-mediated degradation of PPARγ1 in vitro, indicating large MuRF2 increases may lead to PPAR degradation if found in other disease states.Mutations in MuRF2 have been described to contribute to the severity of familial hypertrophic cardiomyopathy.These studies also identify MuRF2 as the first ubiquitin ligase to regulate cardiac PPARα and PPARγ1 activities in vivo via post-translational modification without degradation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, 111 Mason Farm Road, MBRB 2340B, Chapel Hill, NC, USA. heju@email.unc.edu.

ABSTRACT

Background: In diabetes mellitus the morbidity and mortality of cardiovascular disease is increased and represents an important independent mechanism by which heart disease is exacerbated. The pathogenesis of diabetic cardiomyopathy involves the enhanced activation of PPAR transcription factors, including PPARα, and to a lesser degree PPARβ and PPARγ1. How these transcription factors are regulated in the heart is largely unknown. Recent studies have described post-translational ubiquitination of PPARs as ways in which PPAR activity is inhibited in cancer. However, specific mechanisms in the heart have not previously been described. Recent studies have implicated the muscle-specific ubiquitin ligase muscle ring finger-2 (MuRF2) in inhibiting the nuclear transcription factor SRF. Initial studies of MuRF2-/- hearts revealed enhanced PPAR activity, leading to the hypothesis that MuRF2 regulates PPAR activity by post-translational ubiquitination.

Methods: MuRF2-/- mice were challenged with a 26-week 60% fat diet designed to simulate obesity-mediated insulin resistance and diabetic cardiomyopathy. Mice were followed by conscious echocardiography, blood glucose, tissue triglyceride, glycogen levels, immunoblot analysis of intracellular signaling, heart and skeletal muscle morphometrics, and PPARα, PPARβ, and PPARγ1-regulated mRNA expression.

Results: MuRF2 protein levels increase ~20% during the development of diabetic cardiomyopathy induced by high fat diet. Compared to littermate wildtype hearts, MuRF2-/- hearts exhibit an exaggerated diabetic cardiomyopathy, characterized by an early onset systolic dysfunction, larger left ventricular mass, and higher heart weight. MuRF2-/- hearts had significantly increased PPARα- and PPARγ1-regulated gene expression by RT-qPCR, consistent with MuRF2's regulation of these transcription factors in vivo. Mechanistically, MuRF2 mono-ubiquitinated PPARα and PPARγ1 in vitro, consistent with its non-degradatory role in diabetic cardiomyopathy. However, increasing MuRF2:PPARγ1 (>5:1) beyond physiological levels drove poly-ubiquitin-mediated degradation of PPARγ1 in vitro, indicating large MuRF2 increases may lead to PPAR degradation if found in other disease states.

Conclusions: Mutations in MuRF2 have been described to contribute to the severity of familial hypertrophic cardiomyopathy. The present study suggests that the lack of MuRF2, as found in these patients, can result in an exaggerated diabetic cardiomyopathy. These studies also identify MuRF2 as the first ubiquitin ligase to regulate cardiac PPARα and PPARγ1 activities in vivo via post-translational modification without degradation.

No MeSH data available.


Related in: MedlinePlus

Non-targeted cardiac metabolomics of MuRF2−/− mice after 26 weeks challenge with high fat diet. a Principal components analysis, b Partial least squares-discriminant analysis (PLS-DA) and variable importance in the projection analysis, c Heat map of metabolites identified by non-targeted GC/MS analysis of cardiac tissue. d Enrichment by pathway-associated metabolite sets and e location-Based Metabolite sets determined from VIP significant and t test significant metabolites identified. N = 3/group. Significance determined as p < 0.05.
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Fig8: Non-targeted cardiac metabolomics of MuRF2−/− mice after 26 weeks challenge with high fat diet. a Principal components analysis, b Partial least squares-discriminant analysis (PLS-DA) and variable importance in the projection analysis, c Heat map of metabolites identified by non-targeted GC/MS analysis of cardiac tissue. d Enrichment by pathway-associated metabolite sets and e location-Based Metabolite sets determined from VIP significant and t test significant metabolites identified. N = 3/group. Significance determined as p < 0.05.

Mentions: By non-targeted metabolomics analysis, MuRF2 hearts had significant decreases in taurine, myoinositol, and four metabolites involved in malate-aspartate shuttle (glycerol-1phosphate, urea, malic acid, and phosphoric acid) [37]. In the present study, we similarly analyzed MuRF2−/− hearts using non-targeted metabolomics analysis after 26 weeks of high fat diet (Fig. 8). The separation of MuRF2−/− hearts from wildtype was clear using Principal Components Analysis (PCA) (Fig. 8a) as well as Partial Least Squares Discriminant Analysis (PLS-DA) and Variable Interdependent Parameters (VIP) analysis (Fig. 8b). Among all of the annotated metabolites (Fig. 8c), the VIP significant analytes detected were taurine, sucrose, glyceric acid, 3-hydroxyflavone, pantothenic acid, and glutamic acid, among others (Fig. 8b). Enrichment analysis identified the (1) urea cycle; (2) aspartate metabolism; and (3) taurine and hypotaurine metabolism to be the highest fold enriched by metabolite sets (Fig. 8d). Based on location, the mitochondria, peroxisome, and lysosome were the most enriched (Fig. 8e). Pathway analysis identified (1) taurine and Hypotaurine metabolism; (2) glycine, serine, and threonine metabolism; and (3) alanine, aspartate, and glutamate metabolism as the pathways most significantly affected when both t test and VIP significant metabolites were analyzed (Additional file 6: Figure S6).Fig. 8


MuRF2 regulates PPARγ1 activity to protect against diabetic cardiomyopathy and enhance weight gain induced by a high fat diet.

He J, Quintana MT, Sullivan J, L Parry T, J Grevengoed T, Schisler JC, Hill JA, Yates CC, Mapanga RF, Essop MF, Stansfield WE, Bain JR, Newgard CB, Muehlbauer MJ, Han Y, Clarke BA, Willis MS - Cardiovasc Diabetol (2015)

Non-targeted cardiac metabolomics of MuRF2−/− mice after 26 weeks challenge with high fat diet. a Principal components analysis, b Partial least squares-discriminant analysis (PLS-DA) and variable importance in the projection analysis, c Heat map of metabolites identified by non-targeted GC/MS analysis of cardiac tissue. d Enrichment by pathway-associated metabolite sets and e location-Based Metabolite sets determined from VIP significant and t test significant metabolites identified. N = 3/group. Significance determined as p < 0.05.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4526192&req=5

Fig8: Non-targeted cardiac metabolomics of MuRF2−/− mice after 26 weeks challenge with high fat diet. a Principal components analysis, b Partial least squares-discriminant analysis (PLS-DA) and variable importance in the projection analysis, c Heat map of metabolites identified by non-targeted GC/MS analysis of cardiac tissue. d Enrichment by pathway-associated metabolite sets and e location-Based Metabolite sets determined from VIP significant and t test significant metabolites identified. N = 3/group. Significance determined as p < 0.05.
Mentions: By non-targeted metabolomics analysis, MuRF2 hearts had significant decreases in taurine, myoinositol, and four metabolites involved in malate-aspartate shuttle (glycerol-1phosphate, urea, malic acid, and phosphoric acid) [37]. In the present study, we similarly analyzed MuRF2−/− hearts using non-targeted metabolomics analysis after 26 weeks of high fat diet (Fig. 8). The separation of MuRF2−/− hearts from wildtype was clear using Principal Components Analysis (PCA) (Fig. 8a) as well as Partial Least Squares Discriminant Analysis (PLS-DA) and Variable Interdependent Parameters (VIP) analysis (Fig. 8b). Among all of the annotated metabolites (Fig. 8c), the VIP significant analytes detected were taurine, sucrose, glyceric acid, 3-hydroxyflavone, pantothenic acid, and glutamic acid, among others (Fig. 8b). Enrichment analysis identified the (1) urea cycle; (2) aspartate metabolism; and (3) taurine and hypotaurine metabolism to be the highest fold enriched by metabolite sets (Fig. 8d). Based on location, the mitochondria, peroxisome, and lysosome were the most enriched (Fig. 8e). Pathway analysis identified (1) taurine and Hypotaurine metabolism; (2) glycine, serine, and threonine metabolism; and (3) alanine, aspartate, and glutamate metabolism as the pathways most significantly affected when both t test and VIP significant metabolites were analyzed (Additional file 6: Figure S6).Fig. 8

Bottom Line: However, increasing MuRF2:PPARγ1 (>5:1) beyond physiological levels drove poly-ubiquitin-mediated degradation of PPARγ1 in vitro, indicating large MuRF2 increases may lead to PPAR degradation if found in other disease states.Mutations in MuRF2 have been described to contribute to the severity of familial hypertrophic cardiomyopathy.These studies also identify MuRF2 as the first ubiquitin ligase to regulate cardiac PPARα and PPARγ1 activities in vivo via post-translational modification without degradation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, 111 Mason Farm Road, MBRB 2340B, Chapel Hill, NC, USA. heju@email.unc.edu.

ABSTRACT

Background: In diabetes mellitus the morbidity and mortality of cardiovascular disease is increased and represents an important independent mechanism by which heart disease is exacerbated. The pathogenesis of diabetic cardiomyopathy involves the enhanced activation of PPAR transcription factors, including PPARα, and to a lesser degree PPARβ and PPARγ1. How these transcription factors are regulated in the heart is largely unknown. Recent studies have described post-translational ubiquitination of PPARs as ways in which PPAR activity is inhibited in cancer. However, specific mechanisms in the heart have not previously been described. Recent studies have implicated the muscle-specific ubiquitin ligase muscle ring finger-2 (MuRF2) in inhibiting the nuclear transcription factor SRF. Initial studies of MuRF2-/- hearts revealed enhanced PPAR activity, leading to the hypothesis that MuRF2 regulates PPAR activity by post-translational ubiquitination.

Methods: MuRF2-/- mice were challenged with a 26-week 60% fat diet designed to simulate obesity-mediated insulin resistance and diabetic cardiomyopathy. Mice were followed by conscious echocardiography, blood glucose, tissue triglyceride, glycogen levels, immunoblot analysis of intracellular signaling, heart and skeletal muscle morphometrics, and PPARα, PPARβ, and PPARγ1-regulated mRNA expression.

Results: MuRF2 protein levels increase ~20% during the development of diabetic cardiomyopathy induced by high fat diet. Compared to littermate wildtype hearts, MuRF2-/- hearts exhibit an exaggerated diabetic cardiomyopathy, characterized by an early onset systolic dysfunction, larger left ventricular mass, and higher heart weight. MuRF2-/- hearts had significantly increased PPARα- and PPARγ1-regulated gene expression by RT-qPCR, consistent with MuRF2's regulation of these transcription factors in vivo. Mechanistically, MuRF2 mono-ubiquitinated PPARα and PPARγ1 in vitro, consistent with its non-degradatory role in diabetic cardiomyopathy. However, increasing MuRF2:PPARγ1 (>5:1) beyond physiological levels drove poly-ubiquitin-mediated degradation of PPARγ1 in vitro, indicating large MuRF2 increases may lead to PPAR degradation if found in other disease states.

Conclusions: Mutations in MuRF2 have been described to contribute to the severity of familial hypertrophic cardiomyopathy. The present study suggests that the lack of MuRF2, as found in these patients, can result in an exaggerated diabetic cardiomyopathy. These studies also identify MuRF2 as the first ubiquitin ligase to regulate cardiac PPARα and PPARγ1 activities in vivo via post-translational modification without degradation.

No MeSH data available.


Related in: MedlinePlus