The transcriptional regulatory protein HSF1 is the key mediator of induced heat shock protein gene expression in response to elevated temperature and other stresses. Our previous studies identified stress-induced SUMO-1 modification of HSF1 as an important regulator of the DNA-binding activity of this factor. The underlying molecular mechanism by which stress leads to sumoylation of HSF1 was unknown. Prompted by previous studies indicating stress-induced phosphorylation at serine 307 of HSF1, a site very near the sumoylation site at lysine 298, we examined the role of this phosphorylation event in regulating SUMO-1 modification of HSF1. Using a combination of transfection and in vitro phosphorylation/sumoylation experiments, our results indicate that phosphorylation at serine 307 stimulates sumoylation of HSF1. Our results also reveal a role for a conserved leucine zipper sequence in the C-terminal region of HSF1 in inhibiting its SUMO-1 modification. Based on these data, we postulate that phosphorylation at serine 307 could stimulate HSF1 sumoylation by causing a conformation change that relieves the inhibitory effect of the C-terminal leucine zipper. (C) 2003 Elsevier Science (USA). All rights reserved.