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Cyclic strain stimulates isoform‐specific PKC activation and translocation in cultured human keratinocytes
Author(s) -
Takei Teiji,
Han Okhee,
Ikeda Masataka,
Male Philippe,
Mills Ira,
Sumpio Bauer E.
Publication year - 1997
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/(sici)1097-4644(19971201)67:3<327::aid-jcb5>3.0.co;2-y
Subject(s) - protein kinase c , staurosporine , calphostin c , calphostin , microbiology and biotechnology , keratinocyte , cytosol , pkc alpha , gene isoform , biology , signal transduction , chemistry , in vitro , biochemistry , enzyme , gene
Previous studies have demonstrated that cyclic strain induces keratinocyte proliferative and morphological changes. Since protein kinase C (PKC) is known to play an important role in the regulation of keratinocyte growth and differentiation, the objective of this study was to determine the role of the PKC signaling pathway as a mediator of strain modulation of the keratinocyte phenotype. In particular, we tested the following specific hypotheses: (1) cyclic strain stimulates PKC activity and translocation, (2) cyclic strain activates PKC in an isoform‐specific manner, and (3) PKC mediates the strain activated proliferative and morphological response in cultured human keratinocytes. To test these hypotheses, keratinocytes were subjected to vacuum‐generated cyclic strain (10% average strain), followed by measurement of PKC activity, PKC isoform distribution by Western blot analysis and confocal microscopy, and examination of the effect of PKC inhibitors (calphostin C and staurosporine) on strain induced proliferative and morphological changes. We observed stimulation of PKC activity (62.3 ± 5.1% increase) coupled with translocation of PKC from the cytosolic to the membrane fraction in keratinocytes subjected to acute cyclic strain. Cyclic strain also caused translocation of PKC α and δ, but not ζ isoforms, from the cytosolic to the membrane fraction as demonstrated by both Western blot analysis and confocal microscopy. PKC β was not detected in these cells. PKC inhibitors, calphostin C (10 nM), and staurosporine (5 nM), inhibited strain‐induced PKC activation and keratinocyte proliferation, but did not block the effects of strain on cellular morphology or alignment. We conclude that these data support our hypothesis that cyclic strain stimulates PKC activity and translocation in an isoform‐specific manner in cultured human keratinocytes. Moreover, our studies with PKC inhibitors support the hypothesis that strain‐induced changes in the keratinocyte phenotype may be selectively modulated by PKC. J. Cell. Biochem. 67:327–337, 1997. © 1997 Wiley‐Liss, Inc.