Tcn1p/Crz1p, a calcineurin-dependent transcription factor that differentially regulates gene expression inSaccharomyces cerevisiae

Abstract

When fused to the transcription activation domain of Gal4p, the carboxy terminal domain of Tcn1p directed strong calcineurin-independent expression of PMC1-lacZ and other target genes. The amino-terminal domain of Tcn1p was found to function as a calcineurin-dependent transcription activation domain when fused to the DNA-binding domain of Gal4p. This amino-terminal domain also formed Ca 2+-dependent and FK506-sensitive interactions with calcineurin in the yeast two-hybrid assay. These findings suggest that Tcn1p functions as a calcineurin-dependent transcription factor. Interestingly, induction of Tcn1p-dependent genes was found to be differentially controlled in response to physiological Ca 2+ signals generated by treatment with mating pheromone and high salt. We propose that different promoters are sensitive to variations in the strength of Ca 2+ signals generated by these stimuli and to effects of other signaling pathways.

Life in the midst of scarcity: adaptations to nutrient availability in Saccharomyces cerevisiae

“…-ATPase ENA1/PMR2, the Ca 2? -ATPases PMC1 and PMR1 and the catalytic subunit of the b-1-3 glucan synthase FKS2 (Matheos et al 1997;Stathopoulos and Cyert 1997;Stathopoulos-Gerontides et al 1999). Ion stress responses are often transient and after adaptation the subsequent nuclear export of Crz1 occurs via interaction with its export factor Msn5 (Boustany and Cyert 2002).…”

A small molecule‐directed approach to control protein localization and function

“…(D) Schematic illustration of native calcium-responsive signalling pathways (left) and rapamycin-driven Crz1p nuclear localization (right). The results presented here suggest that localization of Crz1p within the nucleus is insufficient to enable target gene expression in the absence of native post-translational modifications and conformational changes conferred by calcium-responsive calcineurin signalling pathways Figure 6B presents a heat map of 68 genes identified as targets of Crz1p in previously published microarray studies (Hilioti et al, 2004;Yoshimoto et al, 2002), including the well-characterized Crz1p targets PMC1, CMK2, GSC2 and RCN1 (Hilioti et al, 2004;Matheos et al, 1997); collectively, the gene set presented here represents the strongest transcriptional profile of Crz1p activity. Upon rapamycin treatment, however, the transcriptional changes observed in these genes are minimal, with only nine genes yielding statistically significant expression changes by significance analysis of microarrays (SAM).…”

“…Specifically, we integrated a vYFP-FRB cassette at the 3 -end of CRZ1 for use with the FKBP-NLS fusion. The Crz1p-vYFP-FRB chimera was functional: the Crz1p fusion exhibited wild-type localization in normal and high-calcium media and, unlike a crz1 null mutant (Matheos et al, 1997), exhibited wild-type growth properties in medium supplemented with 200 mM CaCl 2 (data not shown). In response to elevated CaCl 2 , the Crz1p-vYFP-FRB fusion localized strongly to the nucleus, exhibiting 20-fold nuclear enrichment relative to its localization under conditions of normal calcium ( Figure 6C).…”

“…Towards this end, we applied our drug-induced localization system to control the subcellular distribution of the calcium-regulated transcription factor Crz1p (Matheos et al, 1997;Stathopoulos and Cyert, 1997). In response to transitory rises in cytosolic-free Ca 2+ , Crz1p translocates from the cytoplasm to the nucleus through a complex signalling pathway, outlined schematically in Figure 6.…”

“…Ultimately, these signalling events increase the efficiency of nuclear import of Crz1p, in part through recognition of the Crz1p NLS by the karyopherin Nmd5p (Polizotto and Cyert, 2001), and decrease the efficiency of Crz1p nuclear export (Alepuz et al, 1999;Boustany and Cyert, 2002). Previous studies have suggested that the end purpose of these signalling events is not simply to enable Crz1p translocation to the nucleus, but also to confer modifications onto Crz1p required for its transcriptional activity (Matheos et al, 1997;Boustany and Cyert, 2002). Our platform provides a useful methodology with which to substantiate this model.…”