Mechanical Strain Regulates Contractile Protein Dynamics Related to Heart Remodeling
Swanson, Erik R.
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The mechanisms of sarcomere remodeling in cardiomyocyte hypertrophy are complex, and our previous studies have identified an important role for protein kinase C-ε (PKCε) and the f-actin capping protein CapZ in this process. We now hypothesize that CapZ undergoes PKCε-dependent post-translational modification, which affects its binding kinetics in response to cyclic mechanical strain (CS). Neonatal rat ventricular myocytes were subjected to 10% CS at 1Hz for 1h. CapZ dissociation rates were analyzed by fluorescence recovery after photobleaching (FRAP) using adenoviruses expressing wildtype (wt) GFP-CapZβ1 or wt GFP-CapZβ2. To examine the role of PKCε-dependent post-translational modification of CapZ, cells were infected (100 moi, 24h) with adenoviruses expressing dominant-negative (dn) or constitutively active (ca) PKCε prior to CS. CS enhanced dissociation rates for CapZβ1 (196%, P<0.05) above resting controls, but CapZβ2 was unchanged. caPKCε significantly enhanced dissociation rates for CapZβ1 above controls (176%, P<0.05). Additionally, CapZ post-translational modifications were investigated with quantitative two-dimensional gel electrophoresis (2DGE). 2DGE western blotting identified two post-translational modifications of CapZβ1 and one of CapZβ2. Cyclic strain deceased the unmodified form of CapZβ1 (P<0.05) and increased the doubly modified form (P<0.05) compared to controls, while the addition of dnPKCε with CS reversed this effect (P>0.05). Treatment of caPKCε on unstretched cells increased the singly modified form compared to controls (P<0.05). Neither CS nor either PKCε adenovirus had any effect on CapZβ2 dynamics or post-translational profile. Taken together, our data strongly suggests that CS produces post-translational modification of CapZβ1 but not of CapZβ2. Furthermore, since caPKCε modifies the post-translational profile of CapZβ1 differently from mechanical stress, other mechanisms must also be operative. It seems likely that PKCε post-translational modification and other signaling pathways are both needed for CapZ-actin capping regulation necessary for cell remodeling and hypertrophy.
F-actin capping protein Z
protein kinase C epsilon