Hsf4 counteracts Hsf1 transcription activities and increases lens epithelial cell survival in vitro

The interplay between Hsf4 and Hsf1 plays an important role in the regulation of lens homeostasis. However, the mechanism of the intermolecular association involved is still unclear. In this paper, we find that reconstitution of Hsf4b into Hsf4(-/-) lens epithelial (mLEC/Hsf4(-/-)) cells can simultaneously downregulate Hsp70 expression and upregulate the expression of small heat shock proteins Hsp25 and alpha B-ctystallin at both RNA and protein levels. ChIP assay results indicate Hsf4b, which binds to the promoters of Hsp90 alpha, Hsp70.3, Hsp25 and alpha B-crystallin but not Hsp70.1, can inhibit Hsf1 binding to Hsp70.3 promoter and the heat shock mediated Hsp70 promoter activity by reducing Hsf1 protein expression. Hsf4b N-terminal hydrophobic region can interact with Hsfl N-terminal hydrophobic region. Their interaction impairs Hsf1's intramolecular interaction between the N- and C-terminal hydrophobic regions, leading to Hsf1's cytosolic retention and protein degradation. Both lysosome inhibitors (chloroquine, pepstatin A plus E64d) and proteasome inhibitor MG132 can inhibit Hsf4-mediated Hsfl protein degradation, but MG132 can induce Hsfl activation as well. Upregulation of Hsf4b can significantly inhibit cisplatin and staurosporine induced lens epithelial cell apoptosis through direct upregulation of Hsp25 and alpha B-crystallin expression. Taken together, our results imply that upregulation of Hsf4b modulates the expression pattern of heat shock proteins in lens tissue by either directly binding to their promoters or promoting Hsfl protein degradation. Moreover, upregulation of Hsf4b protects lens cell survival by upregulating anti-apoptotic pathways. These studies reveal a novel regulatory mechanism between Hsfl and Hsf4b in modulating lens epithelial cell homeostasis. (C) 2015 Elsevier B.V. All rights reserved.