Grant: Discovery Grants Program - Individual

The central objective of this research program is to identify and characterize genes and gene products that assist in fungal endophyte colonization of plants. We will use the endophytic, insect pathogenic fungus, Metarhizium as a model system with plants. Here we hypothesize two potential mechanisms where plants allow beneficial fungal interactions. The first objective is to discover novel candidate effector proteins (CEPs) that are effective in allowing fungal colonization of plants. The scientific approach is to: 1) utilize computational tools to identify genes that encode CEPs and use molecular biology techniques to determine their regulation in Metarhizium when symbiotically associated with a host plant; and 2) to perform gene knock-out experiments (with appropriate controls) in order to verify CEPs. These experiments would be complemented with expression of these genes in a yeast model system to determine plant responses to a non-symbiotic fungus. The second objective is to determine the role of small RNAs (sRNAs) in regulating the interactions between Metarhizium and maize plant as the model system. Here the scientific approach is to: 1) discover the composition and expression profiles of sRNAs in Metarhizium during root colonization; 2) to employ computational analysis of maize genes targeted by the up-regulated sRNAs and validated by experimentation; 3) to utilize crosslinking immunoprecipitation, using Argonaute antibodies, to identify the interactions between key sRNAs of Metarhizium and target mRNA of maize genes during root colonization, and 4) to confirm the function of key sRNAs in Metarhizium through overexpression/degradation in Metarhizium and subsequent assessment of the expression of maize target genes together with the ability of Metarhizium to colonize maize roots. For example, sRNAs may target plant mRNA that translate plant proteins for plant immune defence responses. Effector proteins and sRNA are not mutually exclusive mechanisms that allow for fungal colonization of plants but may represent backup or reinforcement mechanisms that are found in many fungal-plant interactions. Using the model insect pathogenic, endophytic fungus Metarhizium the context of this research also has the dimension of exploring the evolutionary processes of Metarhizium, a fungus which is phylogenetically related to plant symbionts. However, during the course of evolution Metarhizium has gained mechanisms to infect insects and can transfer insect-derived nitrogen to plants in exchange for plant carbohydrate. So, there is an added dimension in this research that can be framed in an evolutionary context. Have fungal genes involved in plant symbiosis been appropriated for insect pathogenesis? An understanding of fungal-plant communication is also important in the context of agriculture and accommodating beneficial fungal-plant association and also restricting pathogenic fungi in crop plants.