Complex Haploinsufficiency-Based Genetic Analysis of the NDR/Lats Kinase Cbk1 Provides Insight into Its Multiple Functions in Candida albicans. Issue 3 (1st July 2016)

• Record Type: Journal Article
• Authors: Saputo, Sarah; Norman, Kaitlyn L; Murante, Thomas; Horton, Brooke N; Diaz, Jacinto De La Cruz; DiDone, Louis; Colquhoun, Jennifer; Schroeder, Jeremy W; Simmons, Lyle A; Kumar, Anuj; Krysan, Damian J
• Is Part Of: Genetics. Volume 203:Issue 3(2016)
• Page Start: 1217
• Page End: 1233
• Publication Date: 2016-07-01
• Subjects: C. albicans -- RAM pathway -- complex haploinsufficiency -- fungal morphogenesis; Genetics -- Periodicals; 576.5
• DOI: 10.1534/genetics.116.188029
• Languages: English
• ISSNs: 0016-6731

Abstract

Abstract: Although the analysis of genetic interactions and networks is a powerful approach to understanding biology, it has not been applied widely to the pathogenic yeast Candida albicans . Here, we describe the use of both screening and directed genetic interaction studies based on complex haploinsufficiency to probe the function of the R egulation of A ce2 and M orphogenesis (RAM) pathway in C. albicans . A library of 5200 Tn7 -mutagenized derivatives of a parental strain heterozygous at CBK1, the key kinase in the RAM pathway, was screened for alterations in serum-induced filamentation. Following confirmation of phenotypes and identification of insertion sites by sequencing, a set of 36 unique double heterozygous strains showing complex haploinsufficiency was obtained. In addition to a large set of genes regulated by the RAM transcription factor Ace2, genes related to cell wall biosynthesis, cell cycle, polarity, oxidative stress, and nitrogen utilization were identified. Follow-up analysis led to the first demonstration that the RAM pathway is required for oxidative stress tolerance in a manner related to the two-component-regulated kinase Chk1 and revealed a potential direct connection between the RAM pathway and the essential Mps1 spindle pole-related kinase. In addition, genetic interactions with CDC42 -related genes MSB1, a putative scaffold protein, and RGD3, a putative Rho GTPase-activating protein (GAP) were identified. We also provide evidence that Rgd3 is a GAP for Cdc42 and show that its localization and phosphorylation are dependent on Cbk1.