Protein-Protein Interactions of Protein Kinase C During Polarized Growth in Filamentous Fungi

Rhodes College, Memphis TN

Investigators

Abstract

This project seeks to increase our understanding of how fungi grow. Fungi are microorganisms of great importance to industry, the environment, and agriculture. Many are used to produce commercially valuable products, while others cause major financial losses due to their spoilage of goods and infecting crops. Understanding their growth advances technologies involving their use and aids in developing strategies to control spread of those that are damaging. Fungal cell growth and development are focused at the cell wall, which is an outer cellular structure that provides rigidity and protection from potential environmental stressors. Without a healthy cell wall, fungal growth and the ability to colonize its host are severely compromised. This project builds on previous NSF funded research by the same investigators, which identified candidate proteins involved in fungal cell wall synthesis and growth. In addition to discovering more growth-related proteins, this work will determine which of those proteins physically interact with one another and how they work together during growth. This research will be carried out by a pair of senior scientists working with undergraduate students at Rhodes College (Memphis, TN). Undergraduate students will be integrally involved in all aspects of the research, which will complement and extend their coursework providing a more complete scientific education and better preparing them for advanced studies. The investigators will also continue a program initiated during prior NSF awards, which provides summer research positions to students attending Historically Black Colleges in the Memphis region. This project investigates the function of protein kinase C in filamentous fungal growth and the interplay of its functions with those of other proteins involved in polarized growth and development. Using the filamentous fungus Aspergillus nidulans as a model organism, these researchers and others in the field have identified proteins that localize to sites of cell wall synthesis, namely growing cell apices (hyphal tips) and forming crosswalls (septa). One such protein is PkcA (an A. nidulans homolog of protein kinase C), which is a serine/threonine kinase most similar to the novel protein kinase C isoforms that have been well described in metazoans and yeasts. Although much has been done to describe proteins that participate in growth, this project will add what could be considered the ?PkcA module? to the larger A. nidulans growth network complex. Using techniques including fluorescence labeled proteins, immunoprecipitations, and yeast two-hybrid assays, the researchers will uncover the protein-protein interactions in which PkcA participates at septation sites and hyphal tips and the PkcA motifs responsible for these interactions. In addition, the researchers will determine which complexed proteins are phosphorylated by PkcA using in silico proteomics tools paired with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Not only will this research be of value to the filamentous fungi community, but also to the broader cell biology community, as it will shed light on factors that affect protein recruitment to complexes and how those protein networks function subsequent to recruitment.