Identification and Functional Testing of Novel Interacting Protein Partners for the Stress Sensors Wsc1p and Mid2p of Saccharomyces cerevisiae

G3: Genes, Genomes, Genetics

article

Ednalise Santiago-Cartagena¹  Sahily González-Crespo¹  Vladimir Vélez¹  Nelson Martínez¹  Jamie Snider²  Matthew Jessulat³  Hiroyuki Aoki³  Zoran Minic³  Pearl Akamine¹  Inoushka Mejías¹  Luis M. Pérez¹  Brian C. Rymond⁵  Mohan Babu³  Igor Stagljar²  José R. Rodríguez-Medina¹ 

[1] Department of Biochemistry, University of Puerto Rico, Medical Sciences Campus, PO Box 365067, San Juan, PR 00936-5067;Donnelly Centre, Department of Biochemistry, Department of Molecular Genetics, University of Toronto, Ontario M5S 3E1, Canada;Department of Biochemistry, University of Regina, Regina, Saskatchewan S4S 0A2, Canada;Molecular Sciences and Research Center, University of Puerto Rico, 1390 Ponce de Leon Avenue, Suite 1-7, San Juan, Puerto Rico 00926;Department of Biology, University of Kentucky, Lexington, KY 40506;Mediterranean Institute for Life Sciences, 21000 Split, Croatia

关键词: cell wall;    stress response;    drug resistance;    oxidative stress;    Saccharomyces cerevisiae;   

DOI  :  10.1534/g3.118.200985

学科分类：社会科学、人文和艺术（综合）

来源: Genetics Society of America

【 摘 要 】

Wsc1p and Mid2p are transmembrane signaling proteins of cell wall stress in the budding yeast Saccharomyces cerevisiae . When an environmental stress compromises cell wall integrity, they activate a cell response through the Cell Wall Integrity (CWI) pathway. Studies have shown that the cytoplasmic domain of Wsc1p initiates the CWI signaling cascade by interacting with Rom2p , a Rho1 -GDP-GTP exchange factor. Binding of Rom2p to the cytoplasmic tail of Wsc1p requires dephosphorylation of specific serine residues but the mechanism by which the sensor is dephosphorylated and how it subsequently interacts with Rom2p remains unclear. We hypothesize that Wsc1p and Mid2p must be physically associated with interacting proteins other than Rom2p that facilitate its interaction and regulate the activation of CWI pathway. To address this, a cDNA plasmid library of yeast proteins was expressed in bait strains bearing membrane yeast two-hybrid (MYTH) reporter modules of Wsc1p and Mid2p , and their interacting preys were recovered and sequenced. 14 previously unreported interactors were confirmed for Wsc1p and 29 for Mid2p . The interactors’ functionality were assessed by cell growth assays and CWI pathway activation by western blot analysis of Slt2p / Mpk1p phosphorylation in null mutants of each interactor under defined stress conditions. The susceptibility of these strains to different stresses were tested against antifungal agents and chemicals. This study reports important novel protein interactions of Wsc1p and Mid2p that are associated with the cellular response to oxidative stress induced by Hydrogen Peroxide and cell wall stress induced by Caspofungin.

【 授权许可】

CC BY|CC BY-NC