Targeted Disruption of Transcriptional Regulators inChaetomium globosumActivates Biosynthetic Pathways and Reveals Transcriptional Regulator-Like Behavior of Aureonitol

Abstract

Postgenomic analysis revealed that many microorganisms carry numerous secondary metabolite biosynthetic genes on their genome. However, activities of those putative genes are not clearly reflected in the metabolic profile of the microorganisms, especially in fungi. A recent genome mining effort is promising in discovering new natural products. However, many fungi and other organisms are not amenable to molecular genetics manipulations, making the study difficult. Here we report successful engineering of Chaetomium globosum, a known producer of various valuable natural products, that allows its genetic manipulation via targeted homologous recombination. This strain permitted us to abolish transcriptional regulators associated with epigenetic silencing of secondary metabolite biosynthetic pathways, leading to the identification of the products generated by different gene clusters and isolation of novel secondary metabolites. We were able to identify six gene clusters that are responsible for the biosynthesis of 11 natural products previously known to be produced by C. globosum, including one cytochalasan and six azaphilone-type compounds. In addition, we isolated two new compounds, mollipilin A and B, that were only recently identified in a related Chaetomium species. Furthermore, our investigation into the mechanism of biosynthesis of those natural products in C. globosum also led to the discovery of a secondary metabolite, aureonitol, that acts like a transcriptional regulator for the biosynthesis of other secondary metabolites. Similar approaches should facilitate exploration of the untapped potential of fungal biosynthetic capability and identification of various unique biological functions that those secondary metabolites possess.

Thielavialides A–E, Nor-Spiro-azaphilones, and a Bis-spiro-azaphilone from Thielavia sp. PA0001, An Endophytic Fungus Isolated from Aeroponically Grown Physalis alkekengi

“…Similar oxidative coupling has recently been proposed for the biosynthesis of the related bis-spiro-azaphilone, cochliodone A. 25 …”

Involvement of Lipocalin‐like CghA in Decalin‐Forming Stereoselective Intramolecular [4+2] Cycloaddition

“…We chose to integrate cghA into the chaetoviridin biosynthetic PKS gene cazM [20] so that the integration event would be indicated by the loss of formation of chaetoviridin A 8 and B 9 . [19] A control experiment verified that disruption of cazM would not affect the production of 1 (Figure 4, i vs. ii). Comparison of the extract from the cghA -knockout strain and the cghA -complemented strain showed that the biosynthesis of 7 was no longer observed and exclusive production of 1 was restored with cghA (Figure 4, iii vs. iv), indicating CghA is directly responsible for ensuring the stereoselective formation of 1 and suppressing the nonenzymatic formation of the alternative stereoisomer, 7 .…”

“…[17] Because the cgh gene cluster possessed an aldolase-coding cghB , we proposed CghB to catalyze the aldol condensation of two molecules of pyruvic acid to yield the intermediate 4 , which can then be stereoselectively transaminated to form 5 (Figure 1A). To test this hypothesis, we prepared 5 and its diastereomers chemoenzymatically [18] and used them in feeding experiments with an engineered C. globosum strain, CGKW14 [19] (Supporting Methods). After confirming the loss of production of 1 by ΔcghB /CGKW14 (Figure S14 i vs. iii), we fed 5 to this strain to observe the recovery of the production of 1 (Figure 2).…”

Armochaetoglobins A–J: Cytochalasan Alkaloids from Chaetomium globosum TW1-1, a Fungus Derived from the Terrestrial Arthropod Armadillidium vulgare

“…This information was supported by the HMBC correlations from H[2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] and from H-8 to C-23, as well as the 1 H− 1 H COSY spin system of H 2 -20/H 2 -21/H 2 -22.…”