Abstract
The heart is exquisitely sensitive to mechanical stimuli and adapts to increased demands for work by enlarging the cardiomyocytes. In order to determine links between mechano-transduction mechanisms and hypertrophy, neonatal rat ventricular myocytes (NRVM) were subjected to physiologic strain for analysis of the dynamics of the actin capping protein, CapZ, and its post-translational modifications (PTM). CapZ binding rates were assessed after strain by fluorescence recovery after photobleaching (FRAP) of green fluorescent protein (GFP) expressed by a GFP-CapZβ1 adenovirus. To assess the role of the protein kinase C epsilon isoform (PKCε), rest or cyclic strain were combined with specific PKCε activation by constitutively active PKCε, or by inhibition with dominant negative PKCε (dnPKCε) expression. Significant increases of CapZ FRAP kinetics with strain were blunted by dnPKCε, suggesting that PKCε is involved in mechano-transduction signaling. Similar combinations of strain and PKC regulation in NRVMs were studied by PTM profiles of CapZβ1 using quantitative two-dimensional gel electrophoresis. The significantly increased charge on CapZ seen with mechanical strain was reversed by the addition of dnPKCε. Potential clinical relevance was confirmed in vivo by PTMs of CapZ in the failing heart of one-year old transgenic mice over-expressing PKCε. Furthermore, with strain there was significant PKCε translocation to the Z-disc and co-localization with CapZβ1 or α-actinin, which was quantified on confocal images. A hypothetical model is presented proposing that one destination of the mechanotransduction signaling pathways might be for PTMs of CapZ thereby regulating actin capping and filament assembly.
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Acknowledgments
We thank Allen M. Samarel (Loyola University Chicago Stritch School of Medicine, Maywood, IL) for the gift of the CapZ and PKCε DNA constructs. Normal and transgenic PKCε overexpressing mouse ventricles were kindly provided by Dr. Paul Goldspink from tissue stored at UIC. This work was supported by NIH HL-62426 (BR) and American Heart Association 12PRE12050371 (Y.-H. Lin).
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Ying-Hsi Lin and Erik R. Swanson are co-first authors.
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Lin, YH., Swanson, E.R., Li, J. et al. Cyclic mechanical strain of myocytes modifies CapZβ1 post translationally via PKCε. J Muscle Res Cell Motil 36, 329–337 (2015). https://doi.org/10.1007/s10974-015-9420-6
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DOI: https://doi.org/10.1007/s10974-015-9420-6