Title

Function and localization of the Aspergillus nidulans formin SEPA

Date of Completion

January 2003

Keywords

Biology, Molecular|Biology, Genetics|Biology, Cell

Degree

Ph.D.

Abstract

Cytokinesis, the process by which all cells divide, ensures the appropriate partitioning of nuclear and cytoplasmic contents between daughter cells. The filamentous fungus, Aspergillus nidulans, undergoes cytokinesis by septation, a process in which a chitin crosswall, termed a septum, is synthesized between two hyphal compartments. A genetic screen for mutants defective in septation in A. nidulans identified the gene sepA . Mutants containing sepA1, a temperature-sensitive allele of sepA, fail to form septa at the restrictive temperature. In addition, sepA1 mutants contain wide hyphae and undergo an aberrant branching pattern at their hyphal tips, two phenotypes indicative of defective polarized growth. ^ We have cloned the sepA gene and determined that the sepA gene product encodes a member of the formin family of proteins. Formins are defined by the presence of conserved domains, and are thought to function as molecular scaffolds in actin filament organization. In A. nidulans, actin filaments are required for hyphal tip growth and are required to form the actin ring which precedes septation. To elucidate the role of SEPA in actin dynamics, we have constructed a sepA deletion mutant and determined that sepA is required for actin ring formation. Furthermore, we have localized SEPA to hyphal tips and sites of septation. Using live imaging, we have found that SEPA localizes as a dynamic crescent and spot at hyphal tips, and as a ring that constricts at the sites of septation. Furthermore, we have determined that SEPA co-localizes with actin at both sites. ^ We have also tested the role of various cis and trans-acting factors in SEPA localization. We demonstrate that actin filaments are required to maintain the proper localization pattern of SEPA. In addition, we have determined that the amino-terminal half of SEPA is sufficient for localization to septation sites and hyphal tips. We have also found that the sepH gene product is required for SEPA localization only at septation sites. Our findings support a model in which SEPA directs the formation of the contractile actin ring during septation and functions simultaneously in the formation of actin filament structures required for polarized growth at hyphal tips. ^