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Phytotoxin production in Aspergillus terreus is regulated by independent environmental signals.

Gressler M, Meyer F, Heine D, Hortschansky P, Hertweck C, Brock M - Elife (2015)

Bottom Line: Here, signals, mediators, and biological effects of terrein production were studied in the fungus Aspergillus terreus to elucidate the contribution of terrein to ecological competition.Terrein causes fruit surface lesions and inhibits plant seed germination.Independent signal transduction allows complex sensing of the environment and, combined with its broad spectrum of biological activities, terrein provides a prominent example of adapted secondary metabolite production in response to environmental competition.

View Article: PubMed Central - PubMed

Affiliation: Microbial Biochemistry and Physiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knoell Institute, Jena, Germany.

ABSTRACT
Secondary metabolites have a great potential as pharmaceuticals, but there are only a few examples where regulation of gene cluster expression has been correlated with ecological and physiological relevance for the producer. Here, signals, mediators, and biological effects of terrein production were studied in the fungus Aspergillus terreus to elucidate the contribution of terrein to ecological competition. Terrein causes fruit surface lesions and inhibits plant seed germination. Additionally, terrein is moderately antifungal and reduces ferric iron, thereby supporting growth of A. terreus under iron starvation. In accordance, the lack of nitrogen or iron or elevated methionine levels induced terrein production and was dependent on either the nitrogen response regulators AreA and AtfA or the iron response regulator HapX. Independent signal transduction allows complex sensing of the environment and, combined with its broad spectrum of biological activities, terrein provides a prominent example of adapted secondary metabolite production in response to environmental competition.

No MeSH data available.


Related in: MedlinePlus

High performance liquid chromatography analyses from culture filtrates of SBUG844 wild-type and two independent atfA overexpression mutants (AnPgpdA:atfA; OE 1 and 2).Strains were cultivated under non-inducing conditions for the wild-type. (A) AMM-G100; (B) AMM-G100Gln50. 1 = terrein. Terrein and its side products are detected from the atfA overexpressing strains but not from the wild-type.DOI:http://dx.doi.org/10.7554/eLife.07861.014
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fig4s3: High performance liquid chromatography analyses from culture filtrates of SBUG844 wild-type and two independent atfA overexpression mutants (AnPgpdA:atfA; OE 1 and 2).Strains were cultivated under non-inducing conditions for the wild-type. (A) AMM-G100; (B) AMM-G100Gln50. 1 = terrein. Terrein and its side products are detected from the atfA overexpressing strains but not from the wild-type.DOI:http://dx.doi.org/10.7554/eLife.07861.014

Mentions: In the wild-type the regulators areA and atfA showed a time-dependent increase in gene expression. While the regulator terR was most strongly upregulated after 12 hr, expression of the TerR-controlled terrein biosynthesis genes terA and terB continued to increase after 12 hr and reached 13.3 and 3.2 times the expression level of the actin control gene at 24 hr (Figure 4B). On the contrary, while complemented mutants behaved like the wild-type, the areA and atfA deletions strongly reduced activation of terR and, in turn, the expression of terA and terB. Results from qRT-PCR perfectly coincided with the substantially reduced terrein production rates under nitrogen limitation in these mutants (Figure 4A). AreA recognises the DNA-binding motif HGATAR, and two adjacent binding sites are generally required for transcriptional activation due to dimer formation of AreA monomers (Ravagnani et al., 1997). In this respect, the terR promoter contains two putative AreA binding sites that match the HGATAR consensus (BS1and BS2; positions −59 and −72 relative to the translational start point). Surface plasmon resonance (SPR) analyses with the Aspergillus nidulans AreA DNA-binding domain, which is 91% identical to the respective A. terreus AreA domain, showed that BS1 and BS2 are recognised with high affinity (Figure 4—figure supplement 2). This strengthens the model of a direct involvement of AreA in the activation of terR expression. However, the reduction of terR expression was less pronounced in the ΔareA than in the ΔatfA background (Figure 4B), which indicates that AreA is not the only activator acting on the terR promoter. Therefore, we additionally searched for putative palindromic AtfA/Sko1 binding sites (5′-TKACGTMA-3′) in the promoter regions of the cluster (Proft et al., 2005). Only one hit (TGACGTCA) was identified in the promoter of the structural gene terC. However, if one mismatch is allowed, there is a putative binding site at position −731 relative to the ATG start codon of terR (5´-TGGCGTCA-3´), but it remains speculative whether this binding site is recognised by A. terreus AtfA. Nevertheless, it should be mentioned that even a single half site of the suggested motif could promote transcription factor binding and promoter induction (Proft et al., 2005). We therefore conclude that, although direct evidence for AtfA binding at the terR promoter is lacking, both transcription factors seem to regulate terR expression. In agreement, terR, terA, and terB expression and terrein production showed the strongest decrease in the ΔareAΔatfA double knock-out mutant (Figure 4A). In addition, a constitutive expression of atfA by the gpdA promoter led to increased terrein production (Figure 4—figure supplement 3). This supports our hypothesis of direct involvement of AtfA in terrein cluster regulation.


Phytotoxin production in Aspergillus terreus is regulated by independent environmental signals.

Gressler M, Meyer F, Heine D, Hortschansky P, Hertweck C, Brock M - Elife (2015)

High performance liquid chromatography analyses from culture filtrates of SBUG844 wild-type and two independent atfA overexpression mutants (AnPgpdA:atfA; OE 1 and 2).Strains were cultivated under non-inducing conditions for the wild-type. (A) AMM-G100; (B) AMM-G100Gln50. 1 = terrein. Terrein and its side products are detected from the atfA overexpressing strains but not from the wild-type.DOI:http://dx.doi.org/10.7554/eLife.07861.014
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4528345&req=5

fig4s3: High performance liquid chromatography analyses from culture filtrates of SBUG844 wild-type and two independent atfA overexpression mutants (AnPgpdA:atfA; OE 1 and 2).Strains were cultivated under non-inducing conditions for the wild-type. (A) AMM-G100; (B) AMM-G100Gln50. 1 = terrein. Terrein and its side products are detected from the atfA overexpressing strains but not from the wild-type.DOI:http://dx.doi.org/10.7554/eLife.07861.014
Mentions: In the wild-type the regulators areA and atfA showed a time-dependent increase in gene expression. While the regulator terR was most strongly upregulated after 12 hr, expression of the TerR-controlled terrein biosynthesis genes terA and terB continued to increase after 12 hr and reached 13.3 and 3.2 times the expression level of the actin control gene at 24 hr (Figure 4B). On the contrary, while complemented mutants behaved like the wild-type, the areA and atfA deletions strongly reduced activation of terR and, in turn, the expression of terA and terB. Results from qRT-PCR perfectly coincided with the substantially reduced terrein production rates under nitrogen limitation in these mutants (Figure 4A). AreA recognises the DNA-binding motif HGATAR, and two adjacent binding sites are generally required for transcriptional activation due to dimer formation of AreA monomers (Ravagnani et al., 1997). In this respect, the terR promoter contains two putative AreA binding sites that match the HGATAR consensus (BS1and BS2; positions −59 and −72 relative to the translational start point). Surface plasmon resonance (SPR) analyses with the Aspergillus nidulans AreA DNA-binding domain, which is 91% identical to the respective A. terreus AreA domain, showed that BS1 and BS2 are recognised with high affinity (Figure 4—figure supplement 2). This strengthens the model of a direct involvement of AreA in the activation of terR expression. However, the reduction of terR expression was less pronounced in the ΔareA than in the ΔatfA background (Figure 4B), which indicates that AreA is not the only activator acting on the terR promoter. Therefore, we additionally searched for putative palindromic AtfA/Sko1 binding sites (5′-TKACGTMA-3′) in the promoter regions of the cluster (Proft et al., 2005). Only one hit (TGACGTCA) was identified in the promoter of the structural gene terC. However, if one mismatch is allowed, there is a putative binding site at position −731 relative to the ATG start codon of terR (5´-TGGCGTCA-3´), but it remains speculative whether this binding site is recognised by A. terreus AtfA. Nevertheless, it should be mentioned that even a single half site of the suggested motif could promote transcription factor binding and promoter induction (Proft et al., 2005). We therefore conclude that, although direct evidence for AtfA binding at the terR promoter is lacking, both transcription factors seem to regulate terR expression. In agreement, terR, terA, and terB expression and terrein production showed the strongest decrease in the ΔareAΔatfA double knock-out mutant (Figure 4A). In addition, a constitutive expression of atfA by the gpdA promoter led to increased terrein production (Figure 4—figure supplement 3). This supports our hypothesis of direct involvement of AtfA in terrein cluster regulation.

Bottom Line: Here, signals, mediators, and biological effects of terrein production were studied in the fungus Aspergillus terreus to elucidate the contribution of terrein to ecological competition.Terrein causes fruit surface lesions and inhibits plant seed germination.Independent signal transduction allows complex sensing of the environment and, combined with its broad spectrum of biological activities, terrein provides a prominent example of adapted secondary metabolite production in response to environmental competition.

View Article: PubMed Central - PubMed

Affiliation: Microbial Biochemistry and Physiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knoell Institute, Jena, Germany.

ABSTRACT
Secondary metabolites have a great potential as pharmaceuticals, but there are only a few examples where regulation of gene cluster expression has been correlated with ecological and physiological relevance for the producer. Here, signals, mediators, and biological effects of terrein production were studied in the fungus Aspergillus terreus to elucidate the contribution of terrein to ecological competition. Terrein causes fruit surface lesions and inhibits plant seed germination. Additionally, terrein is moderately antifungal and reduces ferric iron, thereby supporting growth of A. terreus under iron starvation. In accordance, the lack of nitrogen or iron or elevated methionine levels induced terrein production and was dependent on either the nitrogen response regulators AreA and AtfA or the iron response regulator HapX. Independent signal transduction allows complex sensing of the environment and, combined with its broad spectrum of biological activities, terrein provides a prominent example of adapted secondary metabolite production in response to environmental competition.

No MeSH data available.


Related in: MedlinePlus