Discovery of fungal cell wall components using evolutionary and functional genomics

The cell wall is an essential component of fungi that provides shape and protection to cells. As the composition of the wall differentiates pathogenic fungi from the plants and animals they infect, it is a common target for anti-fungal compounds. The goal of the research presented in this dissertation is to make a significant contribution in furthering our understanding of the fungal cell wall and its various components. I have identified and characterized genes involved in cell wall growth and maintenance in the model fungus Neurospora crassa by employing a functional screen based on expression data of vegetative colony development. 14 novel cell wall stress sensitive mutants having multiple phenotypic defects in the categories cell wall morphogenesis, MAPK signaling, Rho-signaling and transport were identified. Taking a broader approach, a combination of phylogenetics, comparative genomic and biochemical approaches was used to extend the understanding of the cell wall components to the early fungal species by reconstructing the phylogenetic history of the fungal cell wall across the evolution of the kingdom. Profile HMMs constructed for proteins involved cell wall biosynthesis, maintenance and remodeling were searched against a collection of fully sequenced genomes to reveal a new class of chitin synthase (CHS) and chitinase (CTS) gene families and the timing of gain of 1,3-beta glucan metabolism machinery in the fungal kingdom. Biochemical characterization of fungal cell walls from early diverging lineages uncovered the existence of noticeably distinct cell walls within the fungal kingdom. The Blastocladiomycota cell walls were found to be similar to Ascomycota i.e. glucan-rich and poor in chitin. Whereas a Chytridomycota fungus had a chitin-rich and glucan-poor cell wall, with mannose being the most substantial component. The Mucormycotina were the most unique having the heteropolymer 'mucoran' as the largest fraction, followed by chitin and very less glucan. This study is the first to report that the Chytridomycete cell walls contain no 1,3 beta-glucan. The presented research should lead to development of better drug targets for designing anti-fungal drugs and enhance our understanding of how cell walls of differing fungal lineages have evolved.