The ability of photodynamic treatment (
PDT) with the
phthalocyanine Pc 4 to activate cellular signal transduction pathways in murine
lymphoma L5178Y-R cells has been assessed by observing increases in
protein tyrosine phosphorylation at early times post-
PDT. Western blot analysis with an anti-
phosphotyrosine antibody revealed a dramatic increase in phosphorylation of two major
protein bands of Mr approximately 80,000 and approximately 55,000 in response to
PDT. The increase was
PDT dose-dependent, occurred as early as 20 s after initiation of light exposure of Pc 4-preloaded cells and was amplified by the presence of the
protein tyrosine phosphatase inhibitor,
sodium orthovanadate (NaVO4). By immunoprecipitation, one of the Mr approximately 80,000 phosphorylated
proteins has been identified as HS1, a substrate of nonreceptor-type
protein tyrosine kinases. Although
vanadate greatly enhanced the level and extent of
PDT-induced phosphorylation, it had no influence on overall photocytotoxicity or on the rate of apoptotic DNA fragmentation.
Genistein, an inhibitor of
protein tyrosine kinases, diminished
tyrosine phosphorylation of the Mr approximately 80,000 and other
proteins and dramatically potentiated cell killing induced by
PDT but did not significantly affect
PDT-induced apoptosis. The results suggest that
PDT rapidly activates a membrane-associated
src family kinase(s) in L5178Y-R cells, one substrate of which is HS1, and that
protein tyrosine phosphorylation is part of a stress response, protecting a portion of the cells from the lethal effects of
PDT but not altering the mechanism by which they die.