Identification of a copper-inducible promoter for use in ectopic expression in the fungal pathogen Histoplasma capsulatum.
Despite the existence of a number of genetic tools to study the fungal pathogen Histoplasma capsulatum, strategies for conditional gene expression have not been developed. We used microarray analysis to identify genes that are transcriptionally induced or repressed by the addition of copper sulfate (CuSO(4)) to H. capsulatum yeast cultures. One of these genes, CRP1, encodes a putative copper efflux pump that is significantly induced in the presence of CuSO(4). The upstream regulatory region of CRP1 was sufficient to drive copper-regulated expression of two reporter genes, lacZ and the gene encoding green fluorescent protein. Microarray experiments were performed to determine a copper concentration that triggers accumulation of the CRP1 transcript without significant perturbation of global gene expression. These studies show that the CRP1 upstream regulatory region can be used for ectopic expression of heterologous genes in H. capsulatum. Furthermore, they demonstrate the strategic use of microarrays to identify conditional promoters that confer induction in the absence of large-scale shifts in gene expression.
Gebhart
Dana
D
Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143-0414, USA.
Bahrami
Adam K
AK
Sil
Anita
A
D000595:Amino Acid Sequence; D019327:Copper Sulfate / Q000494:pharmacology*; D015536:Down-Regulation; D015966:Gene Expression Regulation, Fungal; D017930:Genes, Reporter; D016681:Genome, Fungal; D006658:Histoplasma / Q000235:genetics*; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment; D014170:Transformation, Genetic; D015854:Up-Regulation
D019327:Copper Sulfate