Examination of KNK437- and quercetin-mediated inhibition of heat shock-induced heat shock protein gene expression in Xenopus laevis cultured cells

We examined the effect of quercetin (3,3′,4′,5,7-pentahydroxyflavon) and KNK437 (N-formyl-3,4-methylenedioxy-benzylidene-γ-butyrolactam), a benzylidene lactam compound, on heat-induced heat shock protein (hsp) gene expression in Xenopus laevis A6 kidney epithelial cells. In previous studies, both quercetin and KNK437 inhibited heat shock factor activity resulting in a repression of hsp mRNA and protein accumulation in human cultured cells. In this first study of the effect of these hsp gene expression inhibitors in a non-mammalian cell line, we report that both quercetin and KNK437 reduced the heat shock-induced accumulation of hsp30, hsp47 and hsp70 mRNA in X. laevis cultured cells. However, these inhibitors had no effect on the relative level of a non-heat shock protein mRNA, ef1α, in either control or heat shocked cells. Western blot and immunocytochemical analyses revealed that quercetin partially inhibited HSP30 protein accumulation. In contrast, HSP30 protein was not detectable in KNK437-treated cells. Finally, treatment of A6 cells with KNK437 inhibited the heat shock-induced acquisition of thermotolerance, as determined by preservation of actin filaments and cellular morphology using immunocytochemistry and laser scanning confocal microscopy.