Functional Genes Involved In Mitochondrial Homeostasis And Autophagy In Beauveria Bassiana

Beaueria bassiana is an entomopathogenic fungus with great application potential in biological control.Mitochondria are the power plants of eukaryotic cells and contain their own genomic DNA and double membrane structures.Mitochondrial dynamic is maintained through fusion,division,mitophagy and etc.Autophagy can digest damaged or unnecessary proteins and organelles,and plays very important roles in fungal physiology.In our research,we functionally analyzed the roles of mitochondrion related genes and autophagy related genes in development and virulence of B.bassiana.The functional analysis of Bb Mtf The mitochondrial inner membrane bends inward to form the cristae,greatly enhanced the inner space.Mitofilin is an important factor to maintain the complex structure of the mitochondrial membrane(IM).In B.bassiana,Mitofilin is also localized to mitochondria.In this gene-deleted strain,the cristae were scattered and the cristae junction were missing,while the mitochondria in the wild strains had clear cristae junction,and the cristae were arranged neatly.The absence of Bb Mtf affects the synthesis of ATP which decreased by 33% compared with the wild type.Bb Mtf significantly contributed to fungal asexual development.Conidial production of ?Bb Mtf mutant reduced 40%,and blastospore production declined 70%.On starvation stress plates,the conidial germination was significantly repressed in?Bb Mtf mutant.Finally,disruption of Bb Mtf significantly attenuated fungal virulence against insect hosts.In conclusion,Bb Mtf acts as a mitochondrial protein and is required for maintaining the morphology of the mitochondrial IM,which contributing to energy generation,asexual development and virulence of B.bassiana.The functional analysis of BbFis1,BbMdv1 and BbDnm1 Mitochondrial division is regulated by three genes(Fis1,Mdv1 and Dnm1).In B.bassiana,mutation of Bb Fis1,Bb Mdv1 or Bb Dnm1 resulted in the enlarged mitochondria accumulated in fungal cells.Gene disruption also impaired the energy generation,with reductions ranging from 12% to 16%.On the nutrient-rich SDAY plate and the nutrient-deficient CZA plate with different carbon and nitrogen sources,the colony diameter of the disruption mutant was smaller than those of the wild-type strain.The conidia formation of the gene disruption mutants was delayed,and there was no significant difference between the mutant and the wild-type at 8 d post incubation.The blastospore yield of the mutants was lower than that of the wild type.There mutants displayed the weakened virulence.Additionally,mitophagy was blocked in the ?Bb Fis1,?Bb Mdv1 and?Bb Dnm1 mutant strains.Bb Fis1 was revealed to localize on mitochondria.Together,Bb Fis1,Bb Mdv1 and Bb Dnm1 are all necessary for mitochondrial division which links energy generation and mitophagy to fungal development and virulence in B.bassiana.The functional analysis of Bb Atg6 and Bb Atg16 Autophagy is a highly conserved pathway in eukaryotes.ATG6 and ATG16 both are served as core genes in autophagy,acting on the pre-autophagy and the mid-autophagy respectively.In B.bassiana,Bb Atg6 and Bb Atg16 also are indispensable for autophagy.Gene loss could completely ablate the formation of autophagosomes and autophagic bodies.Bb ATG6 loss changed the cellular distribution of phosphatidylinositol 3-phosphate which indicated with GFP-2×FYVE.Both Bb ATG6 and Bb ATG16 contributed to asexual development of B.bassiana.The blastospore and conidia production of two mutant strains were significantly lower than those in wild type,and Bb ATG16 played a more significant role.Additionally,the loss of Bb ATG6 or Bb ATG16 resulted in the weakened virulence.In conclusion,Bb ATG6 and Bb ATG16 act as important gene in autophagic process,and contribute to asexual development and virulence of B.bassiana.