Calcium induces both
involucrin and
transglutaminase-K in normal keratinocytes (NHK) but not in
squamous carcinoma cell lines (SCC). The
protein kinase C (PKC) agonist
phorbol myristoyl
acetate potentiates and the PKC antagonist
Ro31-8220 blocks the ability of
calcium to stimulate the
involucrin promoter in normal human keratinocytes but not in SCC4. We thus examined the ability of
calcium to regulate the levels of five PKC
isozymes in NHK and two SCC. In the normal keratinocytes, the levels of PKC [alpha], PKC [delta],
PKC [eta], and PKC [zeta] increased over the first one to two weeks in a
calcium-and time-dependent manner.
PKC [epsilon] decreased in a time-and
calcium-dependent fashion over the three-week period. All five
isozymes showed little change during culture in SCC4 at any
calcium concentration.
Calcium and time of culture had partial effects on SCC12B2, a
carcinoma that shows partial differentiation characteristics. Since PKC [alpha] is the only
calcium responsive PKC
isozyme in keratinocytes and most likely to be directly involved in
calcium induced differentiation, we evaluated the effect of inhibiting its production with
antisense oligonucleotides on
calcium-regulated markers of differentiation. We found that the PKC [alpha] specific
antisense oligonucleotide blocked
calcium stimulated
involucrin promoter activity as well as PKC [alpha],
involucrin, and
transglutaminase protein production, whereas the sense
oligonucleotide control did not. We conclude that although a number of PKC
isozymes are regulated during
calcium-induced differentiation, PKC [alpha] plays a necessary role in mediating
calcium-induced differentiation. Failure to regulate PKC [alpha] in SCC4 may underlie at least part of the failure of
calcium to promote differentiation in these cells.