Identification of Mxr1p-binding sites in the promoters of genes encoding dihydroxyacetone synthase and peroxin 8 of the methylotrophic yeast Pichia pastoris

Kranthi, Balla Venkata ; Kumar, Hunsur Rajendra Vinod ; Rangarajan, Pundi N. (2010) Identification of Mxr1p-binding sites in the promoters of genes encoding dihydroxyacetone synthase and peroxin 8 of the methylotrophic yeast Pichia pastoris Yeast, 27 (9). pp. 705-711. ISSN 0749-503X

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Official URL: http://onlinelibrary.wiley.com/doi/10.1002/yea.176...

Related URL: http://dx.doi.org/10.1002/yea.1766

Abstract

Expression of genes involved in methanol metabolism of Pichia pastoris is regulated by Mxr1p, a zinc finger transcription factor. In this study, we studied the target gene specificity of Mxr1p by examining its ability to bind to promoters of genes encoding dihydroxyacetone synthase (DHAS) and peroxin 8 (PEX8), since methanol-inducible expression of these genes is abrogated in mxr1-null mutant strains of P. pastoris. Different regions of DHAS and PEX8 promoter were isolated from P. pastoris genomic DNA and their ability to bind to a recombinant Mxr1p protein containing the N-terminal 150 amino acids, including the zinc finger DNA-binding domain, was examined. These studies reveal that Mxr1p specifically binds to promoter regions containing multiple 5'-CYCC-3' sequences, although all DNA sequences containing the 5'-CYCC-3' motif do not qualify as Mxr1p-binding sites. Key DNA-binding determinants are present outside 5'-CYCC-3' motif and Mxr1p preferably binds to DNA sequences containing 5'-CYCCNY-3' than those containing 5'-CYCCNR-3' sequences. This study provides new insights into the molecular determinants of target gene specificity of Mxr1p, and the methodology described here can be used for mapping Mxr1p-binding sites in other methanol-inducible promoters of P. pastoris.

Item Type:Article
Source:Copyright of this article belongs to John Wiley and Sons.
Keywords:Pichia pastoris; Mxr1p; Methanol Metabolism; Alcohol Oxidase; Dihydroxyacetone Synthase; Peroxin 8
ID Code:57897
Deposited On:30 Aug 2011 07:51
Last Modified:30 Aug 2011 07:51

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