Regulation of yAP-1 nuclear localization in response to oxidative stress

Abstract

The YAP1 gene of Saccharomyces cerevisiae encodes a bZIP‐containing transcription factor that is essential for the normal response of cells to oxidative stress. Under stress conditions, the activity of yAP‐1 is increased, leading to the induced expression of a number of target genes encoding protective enzymes or molecules. We have examined the mechanism of this activation. Upon imposition of oxidative stress, a small increase in the DNA‐binding capacity of yAP‐1 occurs. However, the major change is at the level of nuclear localization; upon induction the yAP‐1 protein relocalizes from the cytoplasm to the nucleus. This regulated localization is mediated by a cysteine‐rich domain (CRD) at the C‐terminus, its removal resulting in constitutive nuclear localization and high level activity. Furthermore, the CRD of yAP‐1 is sufficient to impose regulated nuclear localization of the GAL4 DNA‐binding domain. Amino acid substitutions indicated that three conserved cysteine residues in the CRD are essential for the regulation. We suggest therefore, that these cysteine residues are important in sensing the redox state of the cell and hence regulating yAP‐1 activity.

Crm1 (XpoI) dependent nuclear export of the budding yeast transcription factor yAP‐1 is sensitive to oxidative stress

“…This was not the case for the mutant yAP-1 cm46A5, which has threonine substitutions at cys598 and cys629 and an alanine substitution at cys620. As described previously (Kuge et al 1997), in wild-type cells this mutant is insensitive to oxidative stress and displays a constitutive cytoplasmic localization. A similar observation was made in mcy8 cells grown in methionine-depleted medium.…”

“…We have shown previously (Kuge et al 1997) that the yAP-1 transcription factor, which is critical for the oxidative stress response and resistance to a variety of cytotoxic agents, is regulated at the level of nuclear localization. In unstressed cells it is predominantly cytoplasmic, but it relocates to the nucleus in response to oxidative stress.…”

“…Our previous studies have demonstrated that the regulated localization of yAP-1 was dependent upon the CRD at the C-terminus (Kuge et al 1997). Removal of the CRD resulted in constitutive nuclear localization.…”

“…We have previously shown that the GFP fusion is active and behaves identically to yAP-1 in wild-type cells (Kuge et al 1997). Since yAP-1 protein seems to diffuse easily during fixation (Figs 1D and 3D and Kuge et al 1997), and since over-expression is usually required to detect a weakly abundant protein such as yAP-1 by immunostaining and may cause the deregulation of yAP-1 activity, we employed a GFPyAP-1 fusion to detect yAP-1 localization.…”

“…We have previously shown that the budding yeast transcription factor yAP-1 is also regulated at the level of nucleocytoplasmic transport (Kuge et al 1997). yAP-1 shows homology within its DNA binding domain to members of the mammalian Jun family of proteins (Moye-Rowley et al 1989).…”