Regulation of Morphogenesis and Multicellular Development in Aspergillus
Multicellular development in eukaryotes requires precise integration of a number of fundamental biological processes. Environmental or intrinsic factors can activate diverse signal transduction pathways that elicit tissue- or developmental-specific morphogenesis. Orderly differentiation and spatial organization of diverse cell types frequently requires that regulation of cell-specific gene expression be coupled to alterations in cell cycle regulation. Ultimately, cellular morphogenesis is driven by changes in basic processes controlling cell growth, polarity, and asymmetry in cell division. The long-term goal of this project is to address mechanistic questions concerning multicellular development and tissue formation using the filamentous fungus Aspergillus nidulans. As in higher eukaryotes, external and internal signals induce two highly synchronous developmental events. Asexual reproduction (conidiation) is characterized by the spatiotemporal organization of a limited number of differentiated cell types. Complex transcriptional regulatory interactions coordinate the expression of hundreds of cell-specific genes with critical changes in cell cycle regulation. A conidiation-independent set of regulatory interactions controls both tissue formation during sexual reproduction and cell cycle modifications required during gametogenesis. The objectives of this proposal are directed toward elucidating mechanisms that control multicellular development during conidiation, regulate gametogenesis and sexual tissue development, and integrate these two reproductive processes. Specific objectives are focused upon a molecular analysis of the Medusa (medA), Stunted (stuA) and AnSterile12 (anste12) genes. MedAp is required for commitment to the developmental program, functioning as a co-activator of a subset of conidiation-specific genes and of unknown function in gametogenesis. StuAp is an APSES-domain protein that is an essential component in integrating developmental gene expression and cell cycle regulation. The homeodomain protein AnSte12p is related to fungal proteins known to play essential roles in converting distinct signal transduction inputs into specific developmental events. A balance between functions of StuAp, MedAp and AnSte12p with other developmental regulators establishes cell-specific morphology in Aspergillus nidulans.
During the proposed funding period a series of interrelated classical genetic, molecular genetic and biochemical experiments are designed to: 1) identify DNA response elements within an upstream enhancer required for developmentally regulated, but BrlAp-, AbaAp-independent expression of stuA; 2) begin characterization of novel regulatory factors that bind these response elements; 3) determine if stuA overexpression alone, or in combination with brlA, alters the cell cycle through modification of G1/S and/or G2/M checkpoints; 4) continue characterization of extragenic suppressors of stuA mutations and extragenic mutations that mimic the stunted phenotype; 5) functionally characterize MedAp - is it a novel sequence-specific DNA binding protein?; 6) isolate extragenic mutations that suppress medA -induced growth arrest; 7) physically characterize the A. nidulans ste12 homolog and begin analysis of the biological role of AnSte12p in Aspergillus development. These experiments should provide important insights into similar mechanisms operating in dimorphic fungi as well as regulatory features that distinguish homothallic and heterothallic fungi. These experiments should also provide a greater understanding of the diverse strategies used to direct eukaryotic cell differentiation and responses to environmental factors.