Dataset Information

Complex regulation of PKC?2 and PDK-1/AKT by ROCK2 in diabetic heart.

ABSTRACT:

Objectives

The RhoA/ROCK pathway contributes to diabetic cardiomyopathy in part by promoting the sustained activation of PKC?2 but the details of their interaction are unclear. The purpose of this study was to investigate if over-activation of ROCK in the diabetic heart leads to direct phosphorylation and activation of PKC?2, and to determine if their interaction affects PDK-1/Akt signaling.

Methods

Regulation by ROCK of PKC?2 and related kinases was investigated by Western blotting and co-immunoprecipitation in whole hearts and isolated cardiomyocytes from 12 to 14-week diabetic rats. Direct ROCK2 phosphorylation of PKC?2 was examined in vitro. siRNA silencing was used to confirm role of ROCK2 in PKC?2 phosphorylation in vascular smooth muscle cells cultured in high glucose. Furthermore, the effect of ROCK inhibition on GLUT4 translocation was determined in isolated cardiomyocytes by confocal microscopy.

Results

Expression of ROCK2 and expression and phosphorylation of PKC?2 were increased in diabetic hearts. A physical interaction between the two kinases was demonstrated by reciprocal immunoprecipitation, while ROCK2 directly phosphorylated PKC?2 at T641 in vitro. ROCK2 siRNA in vascular smooth muscle cells or inhibition of ROCK in diabetic hearts reduced PKC?2 T641 phosphorylation, and this was associated with attenuation of PKC?2 activity. PKC?2 also formed a complex with PDK-1 and its target AKT, and ROCK inhibition resulted in upregulation of the phosphorylation of PDK-1 and AKT, and increased translocation of glucose transporter 4 (GLUT4) to the plasma membrane in diabetic hearts.

Conclusion

This study demonstrates that over-activation of ROCK2 contributes to diabetic cardiomyopathy by multiple mechanisms, including direct phosphorylation and activation of PKC?2 and interference with the PDK-1-mediated phosphorylation and activation of AKT and translocation of GLUT4. This suggests that ROCK2 is a critical node in the development of diabetic cardiomyopathy and may be an effective target to improve cardiac function in diabetes.

SUBMITTER: Lin G

PROVIDER: S-EPMC3896488 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Complex regulation of PKCβ2 and PDK-1/AKT by ROCK2 in diabetic heart.

Lin Guorong G Brownsey Roger W RW Macleod Kathleen M KM

PloS one 20140120 1

<h4>Objectives</h4>The RhoA/ROCK pathway contributes to diabetic cardiomyopathy in part by promoting the sustained activation of PKCβ2 but the details of their interaction are unclear. The purpose of this study was to investigate if over-activation of ROCK in the diabetic heart leads to direct phosphorylation and activation of PKCβ2, and to determine if their interaction affects PDK-1/Akt signaling.<h4>Methods</h4>Regulation by ROCK of PKCβ2 and related kinases was investigated by Western blotti ...[more]

PMID: 24466133

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