Characterization of a Penicillium chrysogenum gene encoding a PacC transcription factor and its binding sites in the divergent pcbAB–pcbC promoter of the penicillin biosynthetic cluster

Abstract

Previous work established that pH regulation of gene expression in Aspergillus nidulans, a major determinant of penicillin biosynthesis, is mediated by the zinc-finger transcription factor PacC, an activator of transcription of the isopenicillin N synthase gene. We characterize here the pacC gene from the efficient penicillin producer Penicillium chrysogenum, which functionally complements an A. nidulans pacC null mutation. It encodes a 641-residue polypeptide showing 64% identify to A. nidulans PacC and containing three putative zinc fingers specifically recognizing a 5'-GCCARG-3' hexanucleotide. Penicillium pacC transcript levels are higher under alkaline than under acidic growth conditions and elevate at late stages of growth. The gene contains three PacC-binding sites in its 5'-upstream region. Transcript levels of pcbC (encoding P. chrysogenum isopenicillin N synthase) are low on a repressing carbon source and elevated on a derepressing carbon source. With either carbon source, alkaline pH elevates pcbC transcript levels, correlating with the presence of seven PacC-binding sites in the 1.1 kb pcbAB-pcbC intergenic region and strongly suggesting that pcbC is under direct pacC control. However, in contrast to the situation in A. nidulans, alkaline pH does not override the negative effects of a repressing carbon source, revealing differences in the regulation of the penicillin pathway between Penicillium and Aspergillus.

Alkaline Response Genes of Saccharomyces cerevisiaeand Their Relationship to the RIM101 Pathway

“…In S. cerevisiae, the alkaline-induced transporter Arn4p imports a bacterial siderophore-iron complex (57), thus capitalizing on a bacterial iron acquisition mechanism. In A. nidulans and P. chrysogenum, penicillin synthesis is stimulated at alkaline pH (20,21). ARN4 and the penicillin biosynthesis genes depend upon Rim101p or PacC for expression.…”

“…For example, alkaline pH induces transcription of IPNA and other penicillin biosynthetic genes in Aspergillus nidulans and Penicillium chrysogenum (20,21), the protease gene XPR2 in Yarrowia lipolytica (22), and cell wall protein genes in Candida albicans (23)(24)(25). These responses depend upon a conserved signal transduction pathway (20, 22, 26 -29) that is best understood from work on the A. nidulans transcription factor PacC.…”

Regulation of Gene Expression by Ambient pH in Filamentous Fungi and Yeasts

“…The positive action of PacCmediated pH regulation overrides the negative effects of a repressing carbon source, although pH and carbon regulation normally act in concert, since poor derepressing carbon sources lead to external alkalinization, possibly because they favor the utilization of the carbon skeleton of amino acids with subsequent release of ammonia (42). These findings stimulated the characterization of pacC in the industrially important producer P. chrysogenum (119), whose product also recognizes in vitro the GCCARG PacC consensus (43,122) in the bidirectional promoter of the acvA-ipnA P. chrysogenum homologues pcbAB-pcbC (119). It has been demonstrated that P. chrysogenum pacC mutations equivalent to alkalinity-mimicking pacC c mutations indeed result in penicillin overproduction (T. Suá-rez, H. N. Arst, Jr., G. Turner, and M. A. Peñalva, international patent application PCT/ES00/00464).…”

“…An eye-catching feature is the presence of three perfect copies and one imperfect copy of a 6-residue direct repeat in which the perfect copies contain only acidic and glutamine residues (122). Its significance is unclear but is not likely to be great, since it is not present in isofunctional homologues from organisms as closely related as Aspergillus niger and Penicillium chrysogenum (76,119).…”

Relationship between Secondary Metabolism and Fungal Development

“…At elevated pHs, PacC activates expression of the isopenicillin N synthase gene, which is involved in penicillin biosynthesis. In Penicillium chrysogenum, PacC also activates the expression of the isopenicillin N synthase homologue, pcbC (114). In addition, PacC regulates morphogenesis.…”