The lignin degradative system (LDS) of the white rot basidiomycetous fungus Phanerochaete chrysosporium is capable of degrading the plant cell wall polymer, lignin, and a range of aromatic pollutants. The major components of this nonspecific oxidative system are two extracellular heme-containing peroxidases, manganese peroxidase (MnP) and lignin peroxidase (LiP). The LDS is expressed during secondary metabolic growth, the onset of which is triggered by limiting nutrient nitrogen. Recently, we discovered that MnP expression is also regulated at the level of gene transcription by Mn ion and heat shock and the primary goal of the proposed project is to elucidate on the molecular level the mechanisms of Mn regulation of mnp gene transcription. Isolation and analysis of mutants and titration with promoter copies will be utilized to determine the type of regulation. The promoter regions of P. chrysosporium mnp genes will be analyzed to determine the sequences responsible for Mn and heat shock regulation of transcription, using promoter-reporter constructs, deletion analysis and synthetic promoter elements. DNaseI footprinting and gel mobility shift assays will be used to assay putative DNA- binding transcription factors, and the factors will be isolated. A possible requirement for Mn in post-transcriptional events will be examined.
