Gene expression profiling for the identification of polarity determinants during fungal development: Characterization of NOX enzymes

真核细胞的性发育通常伴随着生殖细胞极性的转变。我们将使用同宗配合的子囊菌大孢囊霉的发育突变体的分子分析与早期繁殖结构的形成块。这些突变体不能进行营养细胞极性的转变，最终导致子囊体的形成，子囊体是性生活周期的第一个细胞结构。使用这些突变体，我们将确定细胞特异性的表达谱相比，子囊孢子线圈菌丝尖端，使用激光显微切割与下一代测序。一个进一步的重点将是对保守的信号通路的组成部分的基因的空间和时间的表达谱。这些分析将通过定位和功能研究完成，包括研究过表达和敲除菌株。当比较G蛋白α亚基基因、nox（NADPH氧化酶）和csn（COP信号体）基因的敲除菌株时，可以观察到显著的表型一致性。这些是否参与决定细胞极性的相同信号传导途径的问题，将通过构建双突变体和三突变体进行进一步的功能分析来解决。

Sexual development in eukaryotic cells is usually accompanied by a shift in cell polarity of generative cells. We will use the homothallic ascomycete Sordaria macrospora for a molecular analysis of developmental mutants with a block in the formation of early propagation structures. These mutants are unable to perform a shift in polarity of vegetative cells that finally results in the formation of ascogonia, the first cellular structures of the sexual life cycle. Using these mutants, we will determine cell-specific expression profiles in ascogonial coils compared to hyphal tips using laser microdissection together with next generation sequencing. A further focus will be on the spatial and temporal expression profiles of genes that are components of conserved signalling pathways. These analyses will be completed by localization and functional studies including the investigation of overexpression and knockout strains. A striking phenotypic coincidence can be observed when knockout strains of G protein alpha subunit genes, nox (NADPH oxidases) and csn (COP signalosome) genes are compared. The question whether these are participating in identical signalling pathways determining cell polarity, will be addressed by construction of double and triple mutants for further functional analysis.