Trichoderma virens produces viridin/viridiol, heptelidic (koningic) acid, several volatile sesquiterpenes and gliotoxin (Q strains) or gliovirin (P strains). We earlier reported that deletion of the terpene cyclase vir4 and a glyceraldehyde-3-phosphate dehydrogenase (GAPDH, designated as vGPD) associated with the “vir” cluster abrogated the biosynthesis of several volatile sesquiterpene metabolites. Here we show that, the deletion of this GAPDH also impairs the biosynthesis of heptelidic acid (a non-volatile sesquiterpene), viridin (steroid) and gliovirin (non-ribosomal peptide), indicating regulation of non-volatile metabolite biosynthesis by this GAPDH that is associated with a secondary metabolism gene cluster. To gain further insights into the details of this novel form of regulation, we identified the terpene cyclase gene responsible for heptelidic acid biosynthesis (hereafter designated as has1) and prove that the expression of this gene is regulated by vGPD. Interestingly, deletion of has1 impaired biosynthesis of heptelidic acid (HA), viridin and gliovirin, but not of volatile sesquiterpenes. Deletion of the vir cluster associated terpene cyclase gene (vir4), located next to the vGPD gene, did not impair biosynthesis of HA, viridin or gliovirin. We thus unveil a novel circuitry of regulation of secondary metabolism where an HA-tolerant GAPDH isoform (vGPD) regulates HA biosynthesis through the transcriptional regulation of the HA-synthase gene (which is not part of the “vir” cluster). Interestingly, impairment of HA biosynthesis leads to the down-regulation of biosynthesis of other non-volatile secondary metabolites, but not of volatile secondary metabolites. We thus provide evidence that the “vir” cluster associated, HA-tolerant GAPDH in T. virens participates in the biosynthesis of volatile sesquiterpenes as a biosynthetic enzyme, and regulates the production of non-volatile metabolites via regulation of HA biosynthesis. The orthologue of the “vir” cluster in Aspergillus oryzae was earlier reported to synthesize HA by another group. Our study thus proves that the same gene cluster can code for unrelated metabolites in different species.

Trichoderma virens产生 viridin/viridiol、heptelidic (koningic) 酸、几种挥发性倍半萜烯和胶质毒素（Q 菌株）或胶质病毒素（P 菌株）。我们先前报道，萜环化酶缺失vir4与“关联和甘油醛-3-磷酸脱氢酶（GAPDH，被指定为VGPD）VIR”簇废除了几种挥发性倍半萜代谢物的生物合成。在这里，我们表明，该 GAPDH 的缺失也会损害庚烷酸（一种非挥发性倍半萜烯）、绿色素（类固醇）和胶质病毒素（非核糖体肽）的生物合成，表明该 GAPDH 对非挥发性代谢物生物合成的调节与次级代谢基因簇相关。为了进一步了解这种新型调控形式的细节，我们鉴定了负责庚烯酸生物合成的萜烯环化酶基因（以下称为has1）并证明该基因的表达受 vGPD 调控。有趣的是，删除has1庚酸 (HA)、绿色素和胶质病毒素的生物合成受损，但挥发性倍半萜烯的生物合成不受影响。位于vGPD基因旁边的vir簇相关萜烯环化酶基因 ( vir4 ) 的缺失不会损害 HA、viridin 或 gliovirin 的生物合成。因此，我们揭示了一种新的次级代谢调节回路，其中耐 HA 的 GAPDH 同种型 (vGPD) 通过 HA 合酶基因（它不是“ vir ”簇的一部分）的转录调节来调节 HA 生物合成。有趣的是，HA 生物合成的受损导致其他非挥发性次级代谢物的生物合成下调，但不会导致挥发性次级代谢物的生物合成下调。因此，我们提供证据表明“ virT. virens中的簇相关、HA 耐受性 GAPDH作为生物合成酶参与挥发性倍半萜的生物合成，并通过调节 HA 生物合成来调节非挥发性代谢物的产生。另一组早先报道了米曲霉中“ vir ”簇的直向同源物合成HA。因此，我们的研究证明，相同的基因簇可以编码不同物种中不相关的代谢物。