Functional Characterization of a PatL Encoding Zn(II)2Cys6 Transcription Factor Involved in Regulation of Grammicin Biosynthesis in Xylaria grammica

다국어 초록 (Multilingual Abstract)

Previous studies have shown that Xylaria grammica KCTC13121BP, an endolichenic fungus, produces grammicin, a secondary metabolite strongly inhibiting Meloidogyne incognita, the parasitic nematode causing root-knot disease. However, it is not known how grammicin biosynthesis is regulated. In this study, we report that XgPatL, a gene encoding a putative Zn(II)2Cys6 DNA transcription factor, functions as a major positive regulator of grammicin biosynthesis. We first established a method for transforming X. grammica using protoplasts. Disruption of XgPatL using this method completely abolished grammicin production, confirming its role in regulating grammicin production. ΔXgpatL also showed significantly reduced mycelial growth and pigmentation. Overexpression of the XgPatL gene using a constitutively expressed promoter, elongation factor 1α (EF1α) and glyceraldehydes-3-phosphate dehydrogenase (GAPD), caused significantly increased grammicin production.

목차 (Table of Contents)

• I. Introduction. 1
• II. Materials and methods . 5
• 1 Fungal isolate and culture conditions 5
• 2. Protoplast preparation 5
• 3. Fungal transformation of deletion mutants and complement. 6
• 4. Southern blot analysis 7
• 5. Overexpression of XgPatL . . 7
• 6. High performance liquid chromatography (HPLC) . 8
• 7. Real-time quantitative reverse transcription PCR. . 9
• III. Result . 12
• 1. Characterizarion of XgPatL 12
• 2. Mycelial expansion and pigmentation 14
• 3. Effect of XgPatL deletion on grammicin production 15
• 4. Deletion of XgPatL significantly decreased the transcription of grammicin biosynthetic genes. 16
• 5. Overexpression of XgPatL . 18
• 6. Grammicin production capacity by overexpression transformants 20
• IV. Discussion . 22
• V. References . 24