Abstract 14923: Disruption of Ca2+ Homeostasis induces activation of Heat Shock Proteins and a Shift in Cytoskeleton Composition
BACKGROUND: Alteration of Ca2+ homeostasis is known to be an important mechanism underlying the progression to heart failure. However, a conditional, cardiac specific SERCA2 knockout (SERCA2 KO) mouse has sustained in vivo cardiac function for several weeks despite dramatically altered cytosolic Ca2+ handling and SR function. So far, it is not clear which biological processes are altered in the SERCA2 KO myocardium in the compensatory phase.
AIM: To examine the molecular mechanism behind this intriguing compensation using a proteomic approach.
METHODS AND RESULTS: Nine days after tamoxifen-induced gene disruption in the SERCA KO mice, the abundance of SERCA2 protein was reduced to 30% of control levels (western blot) yet there were no differences in cardiac function (left atrial diameter, echocardiography, n= 6 for both KO and age matched controls). Using two dimensional gel electrophoresis (pH 4-7 and 6-11) and tandem mass spectrometry, we identified alterations in more than 60 cytosolic proteins. Fundamental changes were found in the cytoskeletal composition with alterations in vinculin, actin, gelsolin and coffilin-2. These were accompanied by regulation of a subset of heat shock proteins (HSPs), HSPB1, alpha-B-crystallin and HSPA5. Knowing that HSPB1 can stabilize proteins comprising the cytoskeleton and myofilament, we focused our experiments on the functional importance of HSPB1. The function of HSPB1 is depending on its phosphorylation status which was increased in the KO animal - including both known and novel sites (S13, S15, S86, T114). In vitro studies in a number of different cell types (HEK293, HL1 and neonatal rat cardiomyocytes), showed that PKC activation (1 μM PMA/1hr) can induce HSPB1 phosphorylation. Finally, we show how the phosphorylation status affects HSPB1 ability to interact with its binding partners.
CONCLUSIONS AND PERSPECTIVES: We have linked altered Ca2+ homeostasis to activation of heat shock proteins and a shift in cytoskeleton composition. Our results show that disruption of Ca2+ homeostasis can give phenotypical effects in the myocardium beyond the contractile apparatus, also involving the cardiomyocyte cellular architecture.
- © 2011 by American Heart Association, Inc.