Analyses of the role of RHO3 and of the WASP-homologue WAL1 for the maintenance of polarized hyphal growth in Candida albicans

rho家族的gtp结合蛋白决定肌动蛋白细胞骨架的组织，从而决定真核细胞的形态发生和细胞形状。极化菌丝生长被认为是人类真菌病原体白色念珠菌的毒力因子。我们可以证明Wiskott-Aldrich综合征蛋白（WASP）的白色念珠菌同源物在这一过程中是必需的。CaWal1p激活的机制目前尚不清楚，特别是因为真菌WASP与人类WASP相比缺乏g蛋白结合结构域。我们之前的工作支持RHO3和WAL1在肌动蛋白细胞骨架的组织和维持极化菌丝生长之间的联系。在提议的项目中，对CaRHO3进行了详细的表征。缺失突变体将被表征，RHO3在二态转换过程中的转录调控将被研究。通过体内延时荧光显微镜测定Rho3-GFP的时空分布。双混合分析用于寻找包含Rho3p和Wal1p的网络的其他组件。这允许在这两种蛋白质之间建立机制联系，并为白色念珠菌皮质肌动蛋白细胞骨架和极化菌丝生长的调节提供见解。

GTP-binding proteins of the Rho-family determine the organization of the actin cytoskeleton and thus morphogenesis and cell shape of eukaryotic cells. Polarized hyphal growth is regarded as a virulence factor of the human fungal pathogen Candida albicans. We could show that the C. albicans homologue of Wiskott-Aldrich Syndrome Proteins (WASP) is required for this process. The mechanism of CaWal1p activation is at present not understood particularly because fungal WASPs lack a G-protein binding domain in contrast to human WASP. Our previous work supports a connection between RHO3 and WAL1 for the organization of the actin cytoskeleton and the maintenance of polarized hyphal growth. In the proposed project a detailed characterization of CaRHO3 is carried out. Deletion mutants will be characterized and the transcriptional regulation of RHO3 during the dimorphic switch will be investigated. Via in vivo time lapse fluorescence microscopy spatial and temporal distribution of Rho3-GFP is determined. Two-hybrid analyses are applied to find additional components of a network that includes Rho3p and Wal1p. This allows to establish a mechanistic connection beween these two proteins and provides insights into the regulation of the cortical actin cytoskeleton and polarized hyphal growth in C. albicans.