Deletion of a Rare Fungal PKS CgPKS11 Promotes Chaetoglobosin A Biosynthesis, Yet Defers the Growth and Development of Chaetomium globosum.

We previously reported that chaetoglobosin A (ChA) exhibits a great potential in the biocontrol of nematodes and pathogenic fungi. To improve the production of ChA, a CRISPR-Cas9 system was created and applied for eliminating potential competitive polyketide products. One of the polyketide synthase encoding genes, Cgpks11, which is putatively involved in the biosynthesis of chaetoglocin A, was disrupted. Cgpks11 deletion led to the overexpression of the CgcheA gene cluster, which is responsible for ChA biosynthesis, and a 1.6-fold increase of ChA. Transcription of pks-1, a melanin PKS, was simultaneously upregulated. Conversely, the transcription of genes for chaetoglocin A biosynthesis, e.g., CHGG_10646 and CHGG_10649, were significantly downregulated. The deletion also led to growth retardation and seriously impaired ascospore development. This study found a novel regulatory means on the biosynthesis of ChA by CgPKS11. CgPKS11 affects chaetoglobosin A biosynthesis, growth, and development in Chaetomium globosum.

A Putative C2H2 Transcription Factor CgTF6, Controlled by CgTF1, Negatively Regulates Chaetoglobosin A Biosynthesis in Chaetomium globosum.

Gα signaling pathway as well as the global regulator LaeA were demonstrated to positively regulate the biosynthesis of chaetoglobosin A (ChA), a promising biotic pesticide produced by Chaetomium globosum. Recently, the regulatory function of Zn2Cys6 binuclear finger transcription factor CgcheR that lies within the ChA biosynthesis gene cluster has been confirmed. However, CgcheR was not merely a pathway specific regulator. In this study, we showed that the homologs gene of CgcheR (designated as Cgtf1) regulate ChA biosynthesis and sporulation in C. globosum NK102. More importantly, RNA-seq profiling demonstrated that 1,388 genes were significant differentially expressed as Cgtf1 deleted. Among them, a putative C2H2 transcription factor, named Cgtf6, showed the highest gene expression variation in zinc-binding proteins encoding genes as Cgtf1 deleted. qRT-PCR analysis confirmed that expression of Cgtf6 was significantly reduced in CgTF1 null mutants. Whereas, deletion of Cgtf6 resulted in the transcriptional activation and consequent increase in the expression of ChA biosynthesis gene cluster and ChA production in C. globosum. These data suggested that CgTF6 probably acted as an end product feedback effector, and interacted with CgTF1 to maintain a tolerable concentration of ChA for cell survival.

A Fungal Diterpene Synthase Is Responsible for Sterol Biosynthesis for Growth.

A conserved open reading frame, dps, is described in Pestalotiopsis microspora, sharing a remarkable similarity with fungal diterpene synthases whose function is less studied. Loss-of-function approach manifested that dps was necessary for the growth and the development of the fungus. A deletion strain, dpsΔ, showed a fundamental retardation in growth, which could deliberately be restored by the addition of exogenous sterols to the media. Gas chromatography-mass spectrometry analysis confirmed the loss of the ability to produce certain sterols. Thus, the tolerance and the resistance of dpsΔ to several stress conditions were impaired. Secondary metabolites, such as the polyketide derivative dibenzodioxocinones, were significantly diminished. At the molecular level, the deletion of dps even affected the expression of genes in the mevalonate pathway. This report adds knowledge about fungal diterpene synthases in Pestalitiopsis microspora.

A Gene Cluster for the Biosynthesis of Dibenzodioxocinons in the Endophyte Pestalotiopsis microspora, a Taxol Producer.

The fungal products dibenzodioxocinones promise a novel class of inhibitors against cholesterol ester transfer protein (CEPT). Knowledge as to their biosynthesis is scarce. In this report, we characterized four more dibenzodioxocinones, which along with a previously described member pestalotiollide B, delimit the dominant spectrum of secondary metabolites in P. microspora. Through mRNA-seq profiling in gα1Δ, a process that halts the production of the dibenzodioxocinones, a gene cluster harboring 21 genes including a polyketide synthase, designated as pks8, was defined. Disruption of genes in the cluster led to loss of the compounds, concluding the anticipated role in the biosynthesis of the chemicals. The biosynthetic route to dibenzodioxocinones was temporarily speculated. This study reveals the genetic basis underlying the biosynthesis of dibenzodioxocinone in fungi, and may facilitate the practice for yield improvement in the drug development arena.