Biological Function Analysis Of Transcription Factors MoMSN2 And MoMYB1 In Magnaporthe Oryzae

Transcription factors (TFs), one of the largest families of trans-acting molecules, play important roles during growth, development and pathogenisis of most fungus. Studying the molecular mechanism of transcription factors in the regulation of growth, development and pathogenicity of M. oryzae will help to find new targets of chemical control and provide new sight for designing comprehensive management of rice blast diseases.ScMsn2 and ScMsn4 encode transcription factors that regulate the general stress response of Saccharomyces cerevisiae. Msn2p and Msn4p regulate genes in response to several stresses, including heat shock, osmotic shock, oxidative stress, low pH, glucose starvation, sorbic acid and high ethanol concentrations, by binding to the STRE element, 5'-CCCCT-3', located in the promoters of these genes. In the first chapter, we cloned the M. oryzae MoMSN2 and characterized its role in conidiation, host cell interactions and virulence. Comparison of the MoMSN2 sequence with those of Saccharomyces cerevisiae Msn2p, Msn4p and Candida glabrata Msn2p, Msn4p revealed a high degree of identity within the zinc-finger domain, but similar to Trichoderma atroviride seb1, it lacks a significant similarity to HD1. Disruption of MoMSN2 in M.oryzae dramatically reduced conidiation. Further investigation revealed that this mutant did not develop any conidiophore. Quantitative real-time RT-PCR (qRT-PCR)-based expression analysis indicated that some homologues of conidiophore-related genes were affected by theΔMomsn2 mutation, included genes MoCOSl, which involved in the early stage of conidiation and mycelial infection and MoMYB1, which predicted to require for initiation of conidiophore development, suggesting that MoMSN2 may function as a transcriptional regulator controlling genes responsible for conidiation. Complementation experiments suggested that MoMSN2 may be a determinant of conidiation. Inoculations of rice intact and wounded leaves with mycelia suggested that MoMSN2 is required for pathogenicity because the deletion mutant cannot form hypha-driven appressoria. When MoCOSl overexpression in theΔMomsn2 mutant, the growth of aerial hyphal and the production of conidiospore can be partially rescued, yeast one-hybrid study showed that MoMSN2 can interact with MoCOSl by binging to its promoter region and we can know that MoMSN2 acts as a transcription factor upstream of the gene MoCOSl. Morphological phenotypes and qRT-PCR of Guy11 andΔMomsn2 revealed that MoMSN2 is required for the pathway of cell wall biosynthesis. Collectively, these results suggest that the downstream targets of M.oryzae MoMSN2 mediate aerial hyphal growth, conidiogenesis, appressorium formation and cell wall biosynthsis.The Myb gene super-family comprises a group of related genes found in plant, animal, and fungal genomes. Myb genes involves in cell proliferation, differentiation, apoptosis. Myb genes can bind specifically to telomeric repeats in vitro and play important role telomeres. In plants, Mybs have been implicated in controlling cell development, cell cycling, respounding to varieties of hormones and envirenmental signals, especially implicated in the processes of second metabolisms (including the anthocyanin pathway). We cloned the M.oryzae MoMYB1 and characterized its role in conidiation, virulence and stress response. The growth rate and conidiation of theΔMomybl mutant were dramatically reduced compared with the wide type Guy11. Further investigation revealed that this mutant did not develop any conidiophore. Quantitative real-time RT-PCR (qRT-PCR)-based expression analysis indicated that some homologues of conidiophore-related genes were affected by theΔMomsn2 mutation, suggesting that MoMYB1 may function as a transcriptional regulator controlling genes responsible for conidiation. TheΔMomyb1 mutant displayed more tolerance to osmotic stressors. Sequence analysis revealed that MoMYB1 putatively encodes a 322-amino-acid Myb protein. Inoculations of rice and barly leaves with mycelia suggested that MoMYB1 is not required for pathogenicity, but it is required for the ablity to form hypha-driven appressoria. Collectively, these results suggest that the downstream targets of MoMYB1 mediate the growth, conidiogenesis, appressorium formation and stress reponse in M.oryzae.