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.
Laser scanning confocal fluorescence microscopy of ascogonial coils which already appear on mycelia after two days of vegetative growth. After 4 days, the formation of young fruiting bodies (protoperithecia) can be observed. The red fluorescence stems from labeling of a developmental protein with the mRFP protein. Excitation, 584 nm; emission, 607 nm.