The purpose of this research is to determine the molecular genetic regulatory mechanisms controlling initiation of developmental pathways in filamentous fungi. We are studying these processes in the filamentous ascomycete Aspergillus nidulans because this organism provides a system for sophisticated genetic and molecular analyses of multicellular development. The work described here is directed at understanding the initial processes controlling differentiation of multicellular asexual reproductive organs, termed conidiophores, that give rise to mitotically derived spores, termed conidia. Over the past several years, many of the basic components in the regulatory pathway controlling conidiophore assembly and conidium production have been described. This has provided the basis for a model outlining the genetic interactions that occur during conidiation. In this model, development is activated upon expression of the brlA gene which encodes a transcriptional regulator and activates expression of other genes required for sporulation. During the previous funding period, we identified and characterized numerous genes that are required for normal activation of the funding period, we identified and characterized numerous genes that are required for normal activation of the brlA gene. The experiments proposed here are aimed at understanding the roles for the products of each of these genes in activating brlA and controlling initiation of conidiophore development. Three projects with interrelated goals are proposed. These are; 1) Detailed characterization of the products of genes identified by mutation (fluG, flbA, flbC, flbD, and other FLB loci) that have been shown to have a critical role in activating development. Our goal is to understand the molecular interactions among the products of these (and other) genes and the mechanisms controlling their activities. 2) Further analysis of genes identified as putative developmental regulator (FAB genes) based on their overexpression phenotypes. We are particularly interested in determining if these genes are required for normal development and how their products interact with genes identified in other ways. 3) Completion of the characterization of the cis- and trans-acting regulatory elements controlling expression of the complex brlA developmental regulatory locus. We propose that the genes described int he first two projects exert their effects on development by directly or indirectly activating brlA expression. This work will contribute to our understanding of the reproductive molecular biology of a group of organisms that includes animal and plant pathogens as well as providing approaches for controlling both the positive and negative activities of fungi. The results of studies proposed here may also apply basic information regarding athe broad questions of how other organisms regulate development and how eukaryotic cells respond to specific signals by activating appropriate developmental pathways.