PAC1 , a pH-Regulatory Gene from Fusarium verticillioides

Abstract

Fumonisins are a group of mycotoxins that contaminate maize and cause leukoencephalomalacia in equine, pulmonary edema in swine, and promote cancer in mice. Fumonisin biosynthesis in Fusarium verticillioides is repressed by nitrogen and alkaline pH. We cloned a PACC-like gene (PAC1) from F. verticillioides. PACC genes encode the major transcriptional regulators of several pH-responsive pathways in other filamentous fungi. In Northern blot analyses, a PAC1 probe hybridized to a 2.2-kb transcript present in F. verticillioides grown at alkaline pH. A mutant of F. verticillioides with a disrupted PAC1 gene had severely impaired growth at alkaline pH. The mutant produced more fumonisin than the wild type when grown on maize kernels and in a synthetic medium buffered at an acidic pH, 4.5. The mutant, but not the wild type, also produced fumonisin B 1 when mycelia were resuspended in medium buffered at an alkaline pH, 8.4. Transcription of FUM1, a gene involved in fumonisin biosynthesis, was correlated with fumonisin production. We conclude that PAC1 is required for growth at alkaline pH and that Pac1 may have a role as a repressor of fumonisin biosynthesis under alkaline conditions.

Low pH regulates the production of deoxynivalenol by Fusarium graminearum

“…At alkaline pH, the transcription factor PacC acts as both an activator of alkali-expressed genes and a repressor of acid-expressed genes through the same consensus promoter recognition motif (Espeso & Arst, 2000;Tilburn et al, 1995). In F. verticillioides, fumonisin production, which occurs at acidic pH, is likely to be negatively regulated by the PacC homologue (Flaherty et al, 2003). In fact, the F. graminearum genome encodes all components of the pH signalling pathway, namely PacC, PalA, PalB, PalC, PalF PalH and PalI (data not shown).…”

“…Upon sensing of an alkaline pH, signalling is activated, resulting in the proteolytic cleavage and activation of PacC, which then upregulates alkaline-responsive genes and represses acid-expressed genes. A functional global pH regulatory system is known to be required for the regulation of sterigmatocystin, penicillin and fumonisin production (Flaherty et al, 2003;Keller et al, 1997a;Shah et al, 1991).…”

Functional characterization of Fusarium verticillioides CPP1, a gene encoding a putative protein phosphatase 2A catalytic subunit

“…Significant progress has been made in elucidating the fumonisin biosynthetic pathway, and regulatory mechanisms associated with the toxin production. The biosynthetic gene cluster, including the polyketide synthase (PKS) gene FUM1 (Proctor et al, 1999(Proctor et al, , 2003Seo et al, 2001), and a number of regulatory genes involved in fumonisin biosynthesis, namely PAC1, FCC1 and ZFR1, have been identified and characterized (Flaherty et al, 2003;Flaherty & Woloshuk, 2004;Shim & Woloshuk, 2001). However, these genes do not show clear epistatic relationships, and therefore it is conceivable that multiple signalling pathways are associated with fumonisin regulation.…”

“…That report suggested that transcriptional regulation of FUM genes can also be affected by putative regulatory gene(s) outside the FUM cluster. Additionally, the complexity of fumonisin regulation is further enhanced by a variety of physiological and nutritional conditions, notably acidic pH and nitrogen stress, which are known to favour or perhaps trigger fumonisin biosynthesis (Flaherty et al, 2003;Shim & Woloshuk, 1999).…”

Identification of a cis-acting factor modulating the transcription of FUM1, a key fumonisin-biosynthetic gene in the fungal maize pathogen Fusarium verticillioides

“…Transcripts from each of the two targets and one reference gene (FUM1, GFP andTUB2) were quantified in each of three independent biological replicates per experimental condition, in analytical triplicates. For standard curves, target sequences from each of the three genes were amplified from genomic DNA of F. verticillioides (primer pairs FUM1_for/_rev, GFP_for/_rev and TUB2_for/_rev in Table S1, the same used for RT-qPCR) ( Flaherty et al, 2003) and cloned in pGEM-T Easy (Promega). Five-point curves were obtained by serially diluting (1:5) the obtained constructs; concentrations ranged from 25 pg to 40 fg of each plasmid for the three target sequences, corresponding to 7.30 × 10 6 -1.17 × 10 4 copies/reaction for FUM1 and to 7.04 × 10 6 -1.13 × 10 4 copies/reaction for GFP and TUB2.…”