There is increasing support for the idea that excessive production of reactive oxygen species (ROS) contributes to the pathogenesis of diabetes. In particular, a strong correlation has been made between increased ROS, activation of specific protein kinase C (PKC) isoforms, and many functional consequences of diabetes. This correlation has been strengthened by recent transgenic experiments in the heart where overexpression of the PKCB2 isoform but not PKCB, decreased cardiac function. Because PKCB2 activity is increased in hearts from diabetic animals, this transgenic model represents a new approach to understand some of the pathogenic mechanisms of diabetic cardiomyopathy. We have developed a novel hypothesis based on data from our laboratory that p90 ribosomal S6 kinase (p90RSK) is a physiological substrate of PKCB2, and that Troponin I (TnI) phosphorylation by p90RSK contributes to decreased cardiac function in diabetes. Data in support of our hypothesis include. 1) TnI phosphorylation inhibits Ca2+-stimulated myosin MgATPase activity, indicating an important role for TnI phosphorylation in contractile function. 2) Phosphorylation of TnI is increased in PKCB2, but not in PKCe transgenic mice. 3) p90RSK activity is increased in myocardial samples from patients with end-stage heart failure. 4) p90RSK activity is increased in myocardial samples from PKCB2, but not PKCe transgenic mice. 5) p90RSK phosphorylates TnI with -ImM substrate affinity. To prove that PKCB2-dependent p90RSK mediated phosphorylation of TnI causes to cardiac contractile dysfunction we propose two aims:
Aim 1 : Define the role of p90RSK in TnI phosphorylation and regulation of Ca2+-stimulated myosin MgATPase activity;
Aim 2 : Determine the role of p90RSK and TnI phosphorylation in cardiac contractile function in vivo.
In Aim 1, we will characterize the sites in TnI phosphorylated by p90RSK using two-dimensional tryptic phosphopeptide mapping, and show that p90RSK phosphorylation of TnI inhibits calcium-dependent myosin MgATPase activity. Next we will show that mutating the TnI residues phosphorylated by p90RSK prevents alterations in MgATPase activity.
In Aim 2, we will generate p90RSK wild-type overexpressing transgenic mice which we expect will exhibit cardiac dysfunction. Then, based on TnI mutation results in Aim 1, we will make a double transgenic that we anticipate will reverse the cardiac dysfunction associated with p90RSK. These results of these experiments will provide a molecular mechanism for the cardiac dysfunction associated with increased PKC activity, based on phosphorylation of TnI by p90RSK.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL066919-03
Application #
6527944
Study Section
Special Emphasis Panel (ZHL1-CSR-Y (S2))
Program Officer
Liang, Isabella Y
Project Start
2000-09-30
Project End
2004-08-31
Budget Start
2002-09-01
Budget End
2003-08-31
Support Year
3
Fiscal Year
2002
Total Cost
$398,750
Indirect Cost
Name
University of Rochester
Department
Internal Medicine/Medicine
Type
Schools of Dentistry
DUNS #
208469486
City
Rochester
State
NY
Country
United States
Zip Code
14627
Shishido, Tetsuro; Woo, Chang-Hoon; Ding, Bo et al. (2008) Effects of MEK5/ERK5 association on small ubiquitin-related modification of ERK5: implications for diabetic ventricular dysfunction after myocardial infarction. Circ Res 102:1416-25
Yan, Chen; Ding, Bo; Shishido, Tetsuro et al. (2007) Activation of extracellular signal-regulated kinase 5 reduces cardiac apoptosis and dysfunction via inhibition of a phosphodiesterase 3A/inducible cAMP early repressor feedback loop. Circ Res 100:510-9
Yan, Chen; Miller, Clint L; Abe, Jun-ichi (2007) Regulation of phosphodiesterase 3 and inducible cAMP early repressor in the heart. Circ Res 100:489-501
Maekawa, Naoya; Abe, Jun-ichi; Shishido, Tetsuro et al. (2006) Inhibiting p90 ribosomal S6 kinase prevents (Na+)-H+ exchanger-mediated cardiac ischemia-reperfusion injury. Circulation 113:2516-23
Itoh, Seigo; Ding, Bo; Shishido, Tetsuro et al. (2006) Role of p90 ribosomal S6 kinase-mediated prorenin-converting enzyme in ischemic and diabetic myocardium. Circulation 113:1787-98
Ding, Bo; Abe, Jun-Ichi; Wei, Heng et al. (2005) A positive feedback loop of phosphodiesterase 3 (PDE3) and inducible cAMP early repressor (ICER) leads to cardiomyocyte apoptosis. Proc Natl Acad Sci U S A 102:14771-6
Itoh, Seigo; Lemay, Serge; Osawa, Masaki et al. (2005) Mitochondrial Dok-4 recruits Src kinase and regulates NF-kappaB activation in endothelial cells. J Biol Chem 280:26383-96
Ding, Bo; Abe, Jun-ichi; Wei, Heng et al. (2005) Functional role of phosphodiesterase 3 in cardiomyocyte apoptosis: implication in heart failure. Circulation 111:2469-76
Itoh, Seigo; Ding, Bo; Bains, Christopher P et al. (2005) Role of p90 ribosomal S6 kinase (p90RSK) in reactive oxygen species and protein kinase C beta (PKC-beta)-mediated cardiac troponin I phosphorylation. J Biol Chem 280:24135-42
Osawa, Masaki; Itoh, Seigo; Ohta, Shinsuke et al. (2004) ERK1/2 associates with the c-Met-binding domain of growth factor receptor-bound protein 2 (Grb2)-associated binder-1 (Gab1): role in ERK1/2 and early growth response factor-1 (Egr-1) nuclear accumulation. J Biol Chem 279:29691-9

Showing the most recent 10 out of 16 publications