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Identification of fat mass and obesity associated (FTO) protein expression in cardiomyocytes: regulation by leptin and its contribution to leptin-induced hypertrophy.

Gan XT, Zhao G, Huang CX, Rowe AC, Purdham DM, Karmazyn M - PLoS ONE (2013)

Bottom Line: The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation.Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects.Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, The University of Western Ontario, London, Ontario, Canada.

ABSTRACT
The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation. Ubiquitously expressed, FTO was identified in heart homogenates although its function is unknown. We studied whether FTO is specifically expressed within the cardiac myocyte and its potential role pertaining to the hypertrophic effect of the adipokine leptin. Most experiments were performed using cultured neonatal rat cardiomyocytes which showed nuclei-specific FTO expression. Leptin significantly increased FTO expression which was associated with myocyte hypertrophy although both events were abrogated by FTO knockdown with siRNA. Administration of a leptin receptor antibody to either normal or obese rats significant reduced myocardial FTO protein expression. Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects. Expression of the cut-like homeobox 1(CUX1) transcriptional factor was significantly increased by leptin although this was restricted to the cathepsin L-dependent, proteolytically-derived shorter p110CUX1 isoform whereas the longer p200CUX1 protein was not significantly affected. Cathepsin L expression and activity were both significantly increased by leptin whereas a cathepsin L peptide inhibitor or siRNA specific for CUX1 completely prevented the leptin-induced increase in FTO expression. The cathepsin L peptide inhibitor or siRNA-induced knockdown of either CUX1 or FTO abrogated the hypertrophic response to leptin. Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent. This study demonstrates leptin-induced FTO upregulation in cardiomyocytes via JAK2/STAT3- dependent CUX1 upregulation and suggests an FTO regulatory function of leptin. It also demonstrates for the first time a functional role of FTO in the cardiomyocyte.

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Regulation of cathepsin L (Cath L) expression and activity by leptin and the role of cathepsin L in leptin-induced FTO upregulation.Panel A shows time-dependent increases in cathepsin L immunofluorescence intensity following leptin (Lep) addition. Control (Ctl) group depicts myocytes not treated with leptin. Panel B provides evidence for leptin-induced increase in cathepsin L activity following leptin administration and its attenuation by the JAK2 inhibitor AG490 (AG) and STAT3 inhibitor S31–201 (S31). RFUs, relative fluorescence units. Panels C and D demonstrate increased gene and protein expression, respectively, of cathespsin L in cardiomyocytes treated with leptin and their prevention by inhibitors of JAK2 and STAT3. For representative Western blots L = leptin. Panels E and F demonstrate prevention of leptin-induced FTO gene and protein upregulation by the cathepsin L peptide inhibitor. Quantitative data are presented as mean ± SEM. N = 7. *P<0.05 from all other groups in panel B and respective control group in panels C to F. +P<0.05 from the leptin alone group in panels C to F. For panels A and B, time refers to hours after leptin addition. Horizontal bar in bottom right images indicates 200 µm.
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pone-0074235-g007: Regulation of cathepsin L (Cath L) expression and activity by leptin and the role of cathepsin L in leptin-induced FTO upregulation.Panel A shows time-dependent increases in cathepsin L immunofluorescence intensity following leptin (Lep) addition. Control (Ctl) group depicts myocytes not treated with leptin. Panel B provides evidence for leptin-induced increase in cathepsin L activity following leptin administration and its attenuation by the JAK2 inhibitor AG490 (AG) and STAT3 inhibitor S31–201 (S31). RFUs, relative fluorescence units. Panels C and D demonstrate increased gene and protein expression, respectively, of cathespsin L in cardiomyocytes treated with leptin and their prevention by inhibitors of JAK2 and STAT3. For representative Western blots L = leptin. Panels E and F demonstrate prevention of leptin-induced FTO gene and protein upregulation by the cathepsin L peptide inhibitor. Quantitative data are presented as mean ± SEM. N = 7. *P<0.05 from all other groups in panel B and respective control group in panels C to F. +P<0.05 from the leptin alone group in panels C to F. For panels A and B, time refers to hours after leptin addition. Horizontal bar in bottom right images indicates 200 µm.

Mentions: The possibility that leptin-induced increase in FTO levels reflected p200CUX1 proteolysis to p110CUX1 was studied by first determining whether leptin affects cathepsin L and then whether a cathespin L inhibitor exerts any influence on leptin-induced FTO elevation. Leptin produced marked and significant increases in cathepsin L gene and protein expression as well as activity which were abrogated by Jak2 and STAT3 inhibition (Figure 7A–7D). Moreover, a peptide cathepsin L inhibitor completely supressed the ability of leptin to increase both FTO gene and protein expression (Figure 7E and 7F).


Identification of fat mass and obesity associated (FTO) protein expression in cardiomyocytes: regulation by leptin and its contribution to leptin-induced hypertrophy.

Gan XT, Zhao G, Huang CX, Rowe AC, Purdham DM, Karmazyn M - PLoS ONE (2013)

Regulation of cathepsin L (Cath L) expression and activity by leptin and the role of cathepsin L in leptin-induced FTO upregulation.Panel A shows time-dependent increases in cathepsin L immunofluorescence intensity following leptin (Lep) addition. Control (Ctl) group depicts myocytes not treated with leptin. Panel B provides evidence for leptin-induced increase in cathepsin L activity following leptin administration and its attenuation by the JAK2 inhibitor AG490 (AG) and STAT3 inhibitor S31–201 (S31). RFUs, relative fluorescence units. Panels C and D demonstrate increased gene and protein expression, respectively, of cathespsin L in cardiomyocytes treated with leptin and their prevention by inhibitors of JAK2 and STAT3. For representative Western blots L = leptin. Panels E and F demonstrate prevention of leptin-induced FTO gene and protein upregulation by the cathepsin L peptide inhibitor. Quantitative data are presented as mean ± SEM. N = 7. *P<0.05 from all other groups in panel B and respective control group in panels C to F. +P<0.05 from the leptin alone group in panels C to F. For panels A and B, time refers to hours after leptin addition. Horizontal bar in bottom right images indicates 200 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3760875&req=5

pone-0074235-g007: Regulation of cathepsin L (Cath L) expression and activity by leptin and the role of cathepsin L in leptin-induced FTO upregulation.Panel A shows time-dependent increases in cathepsin L immunofluorescence intensity following leptin (Lep) addition. Control (Ctl) group depicts myocytes not treated with leptin. Panel B provides evidence for leptin-induced increase in cathepsin L activity following leptin administration and its attenuation by the JAK2 inhibitor AG490 (AG) and STAT3 inhibitor S31–201 (S31). RFUs, relative fluorescence units. Panels C and D demonstrate increased gene and protein expression, respectively, of cathespsin L in cardiomyocytes treated with leptin and their prevention by inhibitors of JAK2 and STAT3. For representative Western blots L = leptin. Panels E and F demonstrate prevention of leptin-induced FTO gene and protein upregulation by the cathepsin L peptide inhibitor. Quantitative data are presented as mean ± SEM. N = 7. *P<0.05 from all other groups in panel B and respective control group in panels C to F. +P<0.05 from the leptin alone group in panels C to F. For panels A and B, time refers to hours after leptin addition. Horizontal bar in bottom right images indicates 200 µm.
Mentions: The possibility that leptin-induced increase in FTO levels reflected p200CUX1 proteolysis to p110CUX1 was studied by first determining whether leptin affects cathepsin L and then whether a cathespin L inhibitor exerts any influence on leptin-induced FTO elevation. Leptin produced marked and significant increases in cathepsin L gene and protein expression as well as activity which were abrogated by Jak2 and STAT3 inhibition (Figure 7A–7D). Moreover, a peptide cathepsin L inhibitor completely supressed the ability of leptin to increase both FTO gene and protein expression (Figure 7E and 7F).

Bottom Line: The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation.Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects.Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, The University of Western Ontario, London, Ontario, Canada.

ABSTRACT
The recently-identified fat mass and obesity-associated (FTO) protein is associated with various physiological functions including energy and body weight regulation. Ubiquitously expressed, FTO was identified in heart homogenates although its function is unknown. We studied whether FTO is specifically expressed within the cardiac myocyte and its potential role pertaining to the hypertrophic effect of the adipokine leptin. Most experiments were performed using cultured neonatal rat cardiomyocytes which showed nuclei-specific FTO expression. Leptin significantly increased FTO expression which was associated with myocyte hypertrophy although both events were abrogated by FTO knockdown with siRNA. Administration of a leptin receptor antibody to either normal or obese rats significant reduced myocardial FTO protein expression. Responses in cardiomyocytes were accompanied by JAK2/STAT3 activation whereas JAK2/STAT3 inhibition abolished these effects. Expression of the cut-like homeobox 1(CUX1) transcriptional factor was significantly increased by leptin although this was restricted to the cathepsin L-dependent, proteolytically-derived shorter p110CUX1 isoform whereas the longer p200CUX1 protein was not significantly affected. Cathepsin L expression and activity were both significantly increased by leptin whereas a cathepsin L peptide inhibitor or siRNA specific for CUX1 completely prevented the leptin-induced increase in FTO expression. The cathepsin L peptide inhibitor or siRNA-induced knockdown of either CUX1 or FTO abrogated the hypertrophic response to leptin. Two other pro-hypertrophic factors, endothelin-1 or angiotensin II had no effect on FTO expression and FTO knockdown did not alter the hypertrophic response to either agent. This study demonstrates leptin-induced FTO upregulation in cardiomyocytes via JAK2/STAT3- dependent CUX1 upregulation and suggests an FTO regulatory function of leptin. It also demonstrates for the first time a functional role of FTO in the cardiomyocyte.

Show MeSH
Related in: MedlinePlus