Dünkler Alexander, Wendland Jürgen
Department of Microbiology, Friedrich Schiller University, Jena, Germany.
Eukaryot Cell. 2007 May;6(5):844-54. doi: 10.1128/EC.00201-06. Epub 2007 Mar 9.
Rho proteins are essential regulators of morphogenesis in eukaryotic cells. In this report, we investigate the role of two previously uncharacterized Rho proteins, encoded by the Candida albicans RHO3 (CaRHO3) and CaCRL1/CaRHO4 genes. The CaRHO3 gene was found to contain one intron. Promoter shutdown experiments using a MET3 promoter-controlled RHO3 revealed a strong cell polarity defect and a partially depolarized actin cytoskeleton. Hyphal growth after promoter shutdown was abolished in rho3 mutants even in the presence of a constitutively active ras1(G13V) allele, and existing germ tubes became swollen. Deletion of C. albicans RHO4 indicated that it is a nonessential gene and that rho4 mutants were phenotypically different from rho3. Two distinct phenotypes of rho4 cells were elongated cell morphology and an unexpected cell separation defect generating chains of cells. Colony morphology of crl1/rho4 resulted in a growth-dependent smooth (long cell cycle length) or wrinkled (short cell cycle length) phenotype. This phenotype was additionally dependent on the rho4 cell separation defect and was also found in a Cacht3 chitinase mutant that shows a strong cytokinesis defect. The overexpression of the endoglucanase encoding the ENG1 gene, but not CHT3, suppressed the cell separation defect of crl1/rho4 but could not suppress the cell elongation phenotype. C. albicans Crl1/Rho4 and Bnr1 both localize to septal sites in yeast and hyphal cells but not to the hyphal tip. Deletion of RHO4 and BNR1 produced similar morphological phenotypes. Based on the localization of Rho4 and on the rho4 mutant phenotype, we propose a model in which Rho4p may function as a regulator of cell polarity, breaking the initial axis of polarity found during early bud growth to promote the construction of a septum.
Rho蛋白是真核细胞形态发生的重要调节因子。在本报告中，我们研究了由白色念珠菌RHO3（CaRHO3）和CaCRL1/CaRHO4基因编码的两种先前未被表征的Rho蛋白的作用。发现CaRHO3基因含有一个内含子。使用MET3启动子控制的RHO3进行的启动子关闭实验显示出强烈的细胞极性缺陷和部分去极化的肌动蛋白细胞骨架。即使在存在组成型活性ras1(G13V)等位基因的情况下，rho3突变体中启动子关闭后的菌丝生长也被消除，并且现有的芽管会肿胀。白色念珠菌RHO4的缺失表明它是一个非必需基因，并且rho4突变体在表型上与rho3不同。rho4细胞的两种不同表型是细胞形态拉长和产生细胞链的意外细胞分离缺陷。crl1/rho4的菌落形态导致生长依赖性的光滑（长细胞周期长度）或皱缩（短细胞周期长度）表型。这种表型还依赖于rho4细胞分离缺陷，并且在显示强烈胞质分裂缺陷 的Cacht3几丁质酶突变体中也发现了这种表型。编码ENG1基因的内切葡聚糖酶的过表达，而不是CHT3，抑制了crl1/rho4的细胞分离缺陷，但不能抑制细胞伸长表型。白色念珠菌Crl1/Rho4和Bnr1都定位于酵母和菌丝细胞的隔膜部位，但不定位于菌丝尖端。RHO4和BNR1的缺失产生了相似的形态学表型。基于Rho4的定位和rho4突变体表型，我们提出了一个模型，其中Rho4p可能作为细胞极性的调节因子，打破早期芽生长期间发现的初始极性轴，以促进隔膜的构建。
