ISSN:
1617-4623
Keywords:
Key words Basidiomycete
;
CT motif
;
Regulation of gene expression
;
Promoter activity
;
S1-sensitive sites
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract The priA gene of the basidiomycete Lentinus edodes possesses a pyrimidine (CT)-rich stretch (26 bp) that includes a short (6-bp) repeat, the elements of which form a mirror repeat at and near the transcriptional initiation sites. A DNA fragment that included this sequence was inserted into pBR322, and the resulting plasmids were introduced into Escherichia coli. Analysis of the susceptibility of these pBR322 derivatives to cleavage by S1 nuclease, following isolation from E. coli, indicated the formation of an open, S1-sensitive structure within and just downstream of the CT/AG-biased sequence. Replacement of two dTMP residues in one of the repeat elements by dGMP resulted in the elimination of the S1-cleavable open structure from the plasmids. To analyze the effect of the CT/AG-biased sequence from priA in the basidiomycete Coprinus cinereus, the integrating vectors pLC2 and pLC2mutCT were used; these contained the wild-type priA promoter and the mutant priA promoter with the aforementioned mutation in the mirror repeat, respectively. The Streptomyces-derived bialaphos resistance gene (bar) was fused downstream of the promoters, and the resulting plasmids, pLC2-bar and pLC2mutCT-bar, were introduced into C. cinereus. Transformants carrying pLC2mutCT-bar grew significantly more slowly on bialaphos-containing agar plates and contained a noticeably lower level of the bar transcript when compared with the transformants obtained with pLC2-bar. These results suggest that an unusual structure induced by the CT/AG-biased sequence is required for efficient gene expression from the priA promoter.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/s004380051167
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