RUI: Elucidating Regulatory Mechanisms in the Diaphanous-related Formin Proteins using an Integrated Approach to Undergraduate Research and Education

Grand Valley State University, Allendale MI

Investigators

Abstract

Intellectual Merit Diaphanous-related formins (DRFs) are a conserved family of Rho GTPase proteins that coordinate the actin and microtubule cytoskeletal networks. Therefore, these proteins influence cellular shape, motility, and cytokinesis; and it is critical that they are regulated and activated only in response to cellular signals. The proteins are autoinhibitory and are normally maintained in an inactive state. DRF proteins are activated by the binding of Rho GTPases. However, recent studies of mammalian DRFs have demonstrated that binding of Rho GTPases alone is insufficient and that another signal may be involved in DRF activation. Although the nature of this additional signal is unknown, preliminary studies indicate that p21-activated kinase 1 (PAK1) specifically phosphorylates the mouse formin mDia2. This project will identify the specific amino acid residues involved in the mDia2-PAK1 binding interaction and determine if PAK1 plays a role in mDia2 regulation by examining the effects of PAK1 phosphorylation of mDia2 on cellular localization, interaction with Rho GTPases, and actin nucleation ability. In addition, this project will identify specific intramolecular interactions involved in the regulation of the DRF proteins, Dishevelled-associated activator of morphogenesis-1 (DAAM1) and mDia3 to understand DAAM1/mDia3 localization, function, and regulation in cells. By specifically characterizing regulatory interactions in three specific members of the DRF family, this research will significantly advance the understanding of cytoskeletal regulation by DRF proteins. Due to the conservation of DRF proteins in organisms as diverse as slime molds, yeast, worms, fruit flies, mice, and humans, as well as the prevalence of the autoinhibitory regulatory mechanism in cellular proteins, the results of the project will be applicable to a broad array of biological systems. Using the knowledge of the specific amino acid residues involved in the regulation of formin proteins, this project will also provide fundamental insight into the cellular localization and cytoskeletal effects of full length DRFs. In addition, the cellular signaling relevance is even more substantial with the potential regulation of DRFs by PAK1, an enzyme known to be involved in a host of relevant cellular processes, such as cell death, cell survival, cell cycle regulation, and neuronal outgrowth. Broader Impact This project will provide research experiences to undergraduate students. The approach of using a combination of biophysical spectroscopy, biochemistry, and cell and molecular biology previously resulted in a publication with undergraduate co-authors. The ability of two undergraduate students to significantly contribute to that study indicates that projects of these types are ideal for cutting-edge undergraduate research. The majority of the current research will be conducted by undergraduates; half of whom will be first generation and/or non-traditional college students. The undergraduate students will be mentored through independent research projects in order to provide comprehensive training in the sciences. As a result of a full-time hypothesis based research experience, the undergraduate students will learn a variety of scientific techniques, increased critical thinking skills, and gain an accurate understanding of experimental science. This will also provide for stronger students in medical and professional schools, as well as in the workplace. While one of the strengths of Grand Valley State University is the significant number of high quality undergraduate students, completion of the RUI projects will provide valuable experiences for even more deserving students and serve to strengthen the research environment of Grand Valley State University. The results generated from this RUI proposal will be disseminated in peer-reviewed journals, as well as presented by undergraduate researchers at local, regional, and national meetings.