Ethanol stress stimulates sumoylation of transcription factor Cst6 which restricts expression of its target genes

Abstract

Sumoylation is an essential post-translational modification that functions in multiple cellular processes, including transcriptional regulation. Indeed, transcription factors represent one of the largest groups of proteins that are modified by the SUMO peptide. Multiple roles have been identified for sumoylation of transcription factors, including regulation of their activity, interaction with chromatin, and binding site selection. Here, we examine how Cst6, a bZIPcontaining sequence-specific transcription factor in Saccharomyces cerevisiae, is regulated by sumoylation. Cst6 is required for survival during ethanol stress and has roles in the utilization of carbon sources other than glucose. We find that Cst6 is sumoylated to appreciable levels in normally growing yeast at Lys residues 139, 461 and 547, and that its sumoylation level increases in ethanol and oxidative stress conditions, but decreases if ethanol is used as the sole carbon source. To understand the role of Cst6 sumoylation during ethanol stress, we generated a yeast strain that expresses a non-sumoylatable mutant form of Cst6. Cellular levels of the mutant protein are moderately reduced compared to the wild-type form, implying that sumoylation promotes Cst6 stability. Although the mutant can bind DNA, chromatin immunoprecipitation (ChIP) analysis shows that its occupancy level is significantly reduced on promoters of some ethanol stress-regulated genes, suggesting that Cst6 recruitment is attenuated or delayed if it can not be sumoylated. Furthermore, impaired Cst6 sumoylation in the mutant strain correlates with elevated expression of some target genes, either constitutively or during induction by ethanol stress. This is most striking for RPS3, which shows dramatically increased expression in the mutant strain. Together, our results suggest that sumoylation controls multiple properties of Cst6 to limit the expression of its target genes. modifications (PTMs) including acetylation, phosphorylation, and ubiquitination (Carr et al. 2015). In recent years, regulation of transcription factors by sumoylation has gained increased attention. Sumoylation, a reversible post-translational modification that plays an essential role in many cellular processes, involves the covalent attachment of a ~12 kDa SUMO (Small Ubiquitin-like Modifier) peptide to specific lysine residues of substrate proteins (Cubeñas-Potts and Matunis 2013; Makhnevych et al. 2009). Similar to ubiquitination, sumoylation is a dynamic process that requires catalysis by three classes of enzymes: activation of SUMO by an E1 enzyme, conjugation to target proteins by an E2 enzyme (Ubc9), and facilitation of transfer by E3 ligases (Flotho and Melchior 2013). In contrast to ubiquitination, in which E3 ubiquitin ligases are the primary determinants of substrate specificity, Ubc9 can directly specify its targets via the SUMO site consensus sequence ΨKxD/E, where Ψ is a hydrophobic residue, K is the lysine to be modified, x is any amino acid, and D/E represents an acidic residue (Dye and Schulma...

Predictive Values of Homeobox Gene A-Antisense Transcript 3 (HOXA-AS3), Cystatin 6 (CST6), and Chromobox Homolog 4 (CBX4) Expressions in Cancer Tissues for Recurrence of Early Colon Cancer After Surgery

“…CST6 has a reduced expression in osteoblastic breast cancer samples and cells and can inhibit tumor infiltration and bone metastasis. 10 In addition to the inhibitory effect, CST6 also exerts a promoting effect on tumors. 11 For instance, CST6 has a significantly elevated expression in triple-negative breast cancer, which is considered to be a tumor-promoting molecule and has correlations with the low disease-free survival rate and high risk of lymph node metastasis.…”