Neurospora crassa Female Development Requires the PACC and Other Signal Transduction Pathways, Transcription Factors, Chromatin Remodeling, Cell-To-Cell Fusion, and Autophagy

Using a screening protocol we have identified 68 genes that are required for female development in the filamentous fungus Neurospora crassa. We find that we can divide these genes into five general groups: 1) Genes encoding components of the PACC signal transduction pathway, 2) Other signal transduction pathway genes, including genes from the three N. crassa MAP kinase pathways, 3) Transcriptional factor genes, 4) Autophagy genes, and 5) Other miscellaneous genes. Complementation and RIP studies verified that these genes are needed for the formation of the female mating structure, the protoperithecium, and for the maturation of a fertilized protoperithecium into a perithecium. Perithecia grafting experiments demonstrate that the autophagy genes and the cell-to-cell fusion genes (the MAK-1 and MAK-2 pathway genes) are needed for the mobilization and movement of nutrients from an established vegetative hyphal network into the developing protoperithecium. Deletion mutants for the PACC pathway genes palA, palB, palC, palF, palH, and pacC were found to be defective in two aspects of female development. First, they were unable to initiate female development on synthetic crossing medium. However, they could form protoperithecia when grown on cellophane, on corn meal agar, or in response to the presence of nearby perithecia. Second, fertilized perithecia from PACC pathway mutants were unable to produce asci and complete female development. Protein localization experiments with a GFP-tagged PALA construct showed that PALA was localized in a peripheral punctate pattern, consistent with a signaling center associated with the ESCRT complex. The N. crassa PACC signal transduction pathway appears to be similar to the PacC/Rim101 pathway previously characterized in Aspergillus nidulans and Saccharomyces cerevisiae. In N. crassa the pathway plays a key role in regulating female development.