Please use this identifier to cite or link to this item: http://scholarbank.nus.edu.sg/handle/10635/29863
Title: ROLES OF A CORE SEPTIN PROTEIN CDC3 AND A SEPTIN ASSOCIATED PROTEIN NAP1 IN CANDIDA ALBICANS POLARIZED GROWTH AND CELL CYCLE
Authors: ZHAO PAN
Keywords: Cdc3, septins, septin ring, cell cycle, phosphorylation, Nap1
Issue Date: 29-Jul-2011
Source: ZHAO PAN (2011-07-29). ROLES OF A CORE SEPTIN PROTEIN CDC3 AND A SEPTIN ASSOCIATED PROTEIN NAP1 IN CANDIDA ALBICANS POLARIZED GROWTH AND CELL CYCLE. ScholarBank@NUS Repository.
Abstract: Septins play important roles in morphogenesis, cell cycle progression and cytokinesis in yeast. Their organization and function are regulated in a cell cycle dependent manner. However, the mechanisms of control remain unclear. This thesis consists of two projects on the study of one septin Cdc3 and a septin associated protein Nap1 in the fungus Candida albicans. The first project is on a septin protein Cdc3. I uncovered that the Candida albicans septin Cdc3 undergoes cell cycle dependent phosphorylation and that phosphorylation at a single amino acid Ser422 is critical for Cdc3 functions. Hyperphosphorylated isoforms are detected in early G1 followed by a period of dephosphorylation; and after the START, Cdc3 phosphorylation increases gradually through the rest of the cell cycle. Phospho-mapping by mass spectrometry identified phosphorylation on S422 near the C terminus. The phosphomimetic S422D mutation causes disorganization of septin structures, severe cytokinetic defects, dramatic cell elongation, and inability of Cdc3 to localize to the neck. In contrast, the nonphosphorylatable S422A mutation produces a much weaker phenotype. Co-immunoprecipitation experiments demonstrate that the S422D mutation greatly weakens Cdc11¿s association with the septin complex and causes premature dissociation of Cdc11 from the ring. The Nim1 kinase Gin4 is involved in the phosphorylation of Cdc3, but the Cdk Cdc28 is not. These findings reveal that controlling the phospho-regulation at a single residue S422 on Cdc3 may play a crucial role in regulating septin assembly/disassembly and stability. The second project focuses on a septin-associated protein Nap1. I report that deletion of C. albicans NAP1 leads to constitutive filamentous growth, higher sensitivity to hyphal induction, and defective septin organization. FCF, a compound known to stabilize septin filaments, can rescue the defects caused by NAP1 deletion. Fluorescence recovery after photo-bleaching (FRAP) analysis of Cdc3-GFP uncovers a more dynamic septin ring in nap1¿ compared to that in WT. In nap1¿ cells, Cdc3 deposits randomly on the cell cortex as spots or partial rings and experiences impairment of phosphorylation. Double deletion of NAP1 and CDC10, another septin protein, results in exacerbated temperature sensitivity, defective septin ring formation and scattered Cdc3 localization. Phospho-mapping by mass spectrometry identified phosphorylation on 10 Thr/Ser residues in the N-terminus of Nap1. Mutation of these 10 residues to non-phosphorylatable Ala results in pseudohyphal growth and affects Nap1 neck localization. Conversely, mutation of these ten residues to phospho-mimetic Glu does not affect cell morphology, but causes random deposition of Cdc3. My findings unveil the roles of Nap1 in septin stabilization and Cdc3 phosphorylation control.
URI: http://scholarbank.nus.edu.sg/handle/10635/29863
Appears in Collections:Ph.D Theses (Open)

Show full item record
Files in This Item:
File Description SizeFormatAccess SettingsVersion 
ZhaoP.pdf2.32 MBAdobe PDF

OPEN

NoneView/Download

Page view(s)

244
checked on Dec 11, 2017

Download(s)

21
checked on Dec 11, 2017

Google ScholarTM

Check


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.