Project description:A spontaneously phenotypically degenerated strain of M. robertsii strain ARSEF 2575 (M. robertsii lc2575; lc = low conidiation) showed a reduction in conidiation and fungal virulence after successive subculturing on artificial medium. However, the conidial production and fungal virulence of a phenotypically degenerated M. robertsii were recovered by serially passaging through a plant host. The DNA methylation level of phenotypically degenerated Metarhizium robertsii M. robertsii lc2575 and this fungi after solider bean passages were tested through the whole genome bisulfite sequencing. The results showed that approximately 0.379 % of cytosines are methylated in the fungi after bean passages, almost the same as the DNA methylation level in M. robertsii lc2575 (0.375%). The distribution of different methylated regions located more on intergenic regions of fungi after bean passages than M. robertsii lc2575. Gene Ontology (GO) analysis and KEGG analysis of DMR-associated genes revealed that amino acid, carbohydrate and fatty acid metabolism.
Project description:Metarhizium robertsii is one of the model fungus species widely used studying insect-fungus interactions. Our previous studies have shown the accumulation of lipid droplets (LDs) play an essential role in generation of cellular turgor pressure to assist fungal penetration of insect cuticles, and autophagy-related proteins are connected with LD biogenesis and cellular accumulation in M. robertsii. However, full proteomes of LDs are unclear in terms of the species of proteins involved in lipid biosynthesis and degradation. We performed experiments by growing the fungi in different nutrient conditions, isolated the LDs and extracted the LD proteins from the cultures after being grown in a nutrient-rich medium (i.e. Sabouraud dextrose broth, SDB), a minimum medium without nitrogen source (MM-N) and MM-N supplemented with oleate. The protein samples were then subject to LC-MS/MS proteome profilings for identifying and postulating the proteins/pathways involved in lipid metabolisms.
Project description:Exopolysaccharide galactosaminogalactan (GAG) is a fungal cell wall component composed of α-1,4 linked galactose, N-acetyl galactosamine and galactosamine, which has been demonstrated in Aspergillus fumigatus in association with fungal adhesion, biofilm formation and virulence. The gene cluster responsible for GAG biosynthesis has only been characterized in Aspergillus fungi. We found that the highly conserved gene cluster for GAG biosynthesis is also present in the insect pathogenic fungi Metarhizium species. Functional investigations in M. robertsii revealed that GAG is only produced on fungal cell wall during fungal germination, filamentation and the formation of the infection structure appressoria. Gene deletions revealed that, relative to the wild-type (WT), the appressorial mucilage production was abolished in the null mutant of M. robertsii. Since multiple enzymes are produced in appressorial mucilages, appressorial samples of the WT and mutant formed on cicada wings were collected and subjected to iTRAQ comparative proteomic analysis. We found that different protein families were up- or down-regulated in the null mutant when compared with the WT.