Novel Promoter Sequence Required for Manganese Regulation of Manganese Peroxidase Isozyme 1 Gene Expression in Phanerochaete chrysosporium

Abstract

ABSTRACT Manganese peroxidase (MnP) is a major, extracellular component of the lignin-degrading system produced by the wood-rotting basidiomycetous fungus Phanerochaete chrysosporium . The transcription of MnP-encoding genes ( mnp s) in P. chrysosporium occurs as a secondary metabolic event, triggered by nutrient-nitrogen limitation. In addition, mnp expression occurs only under Mn 2+ supplementation. Using a reporter system based on the enhanced green fluorescent protein gene ( egfp ), we have characterized the P. chrysosporium mnp1 promoter by examining the effects of deletion, replacement, and translocation mutations on mnp1 promoter-directed egfp expression. The 1,528-bp mnp1 promoter fragment drives egfp expression only under Mn 2+ -sufficient, nitrogen-limiting conditions, as required for endogenous MnP production. However, deletion of a 48-bp fragment, residing 521 bp upstream of the translation start codon in the mnp1 promoter, or replacement of this fragment with an unrelated sequence resulted in egfp expression under nitrogen limitation, both in the absence and presence of exogenous Mn 2+ . Translocation of the 48-bp fragment to a site 120 bp downstream of its original location resulted in Mn 2+ -dependent egfp expression under conditions similar to those observed with the wild-type mnp1 promoter. These results suggest that the 48-bp fragment contains at least one Mn 2+ -responsive cis element. Additional promoter-deletion experiments suggested that the Mn 2+ element(s) is located within the 33-bp sequence at the 3′ end of the 48-bp fragment. This is the first promoter sequence containing a Mn 2+ -responsive element(s) to be characterized in any eukaryotic organism.