| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2005: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2004: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Research Abstract |
A-factor (2-isocapryloyl-3R-hydroxymethyl-γ-butyrolactone) serves as a microbial hormone that induces morphological development and secondary metabolism in Streptomyces griseus. One of the secondary metabolites under the control of A-factor is grixazone (GX) which is a diffusible yellow pigment containing a phenoxazinone chromophore. GX production is also induced by phosphate depletion. Because of the regulation of the GX production by both A-factor and phosphate, the biosynthetic gene cluster for GX is a useful material for elucidating the molecular mechanisms by which γ-butyrolactone and phosphate control secondary metabolite formation.
The biosynthetic gene cluster for GX consists of 13 genes and there are six transcriptional units : griT, griSR, griR, griAB, griCDEFG, and griJIH. Disruption of griR, which encodes a SARP-family transcriptional regulator, abolished the transcription of other genes in the cluster. Recombinant GriR bound to the griC and griJ promoters in vitro. Transcription of griR was not activated in the absence of A-factor or in the presence of a high concentration of phosphate. When griR was transcribed ectopically from a constitutively active promoter, GX was overproduced without A-factor and phosphate-depletion signals. These results indicated that both A-factor and phosphate-depletion signals are transmitted to the GX biosynthesis genes via GriR.
On the other hand, griZ was cloned as a mutated gene in S. griseus M31, a mutant strain deficient in the GX production. griZ exists far away from the GX biosynthetic gene cluster on the chromosome and encodes a TetR-family transcriptional regulator. The griZ-disrupted strain failed to activate griR and to produce GX. When griR was transcribed from a constitutively active promoter in the griZ-disrupted strain, GX production was restored. These results indicated that griZ is essential for the GX production and involved in the activation of griR transcription.
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