Project description:Global transcriptional profile of Streptomyces avermitilis wild-type strain ATCC31267 and avermectin overproducing strain 76-02-e at different time points
Project description:Streptomyces avermitilis is a avermectin producer.Since the avermectin biosynthesis rate has a increased significantly in P3 fermentation stage( P1,24–96 h; P2:96–192 h, P3:192–240 h), but the sugar absorption rate decreased significantly in P3 fermentation stage, in order to improve the titer of avermectins, we conducted transcriptomic analysis of Streptomyces avermitilis S0 in fourth and eighth day, and selected native promoters with appropriate temple using to express sugar transporters.
Project description:The gram-positive bacterium, Streptomyces avermitilis holds industrial importance, which produces widely used anthelmintic agent, avermectin. Furthermore, S. avermitilis is generally considered as a prominent heterologous gene expression host for diverse secondary metabolites biosynthesis. However, despite of its industrial importance, it largely remains unknown how its genome is organized and regulated for timely gene expression. Here, we determined 1,601 transcription units (TU) encoded in its genome using the integrated analysis of high-throughput sequencing data including dRNA-Seq, Term-Seq, RNA-Seq, and Ribo-Seq. In addition to TU cataloguing, these information-rich results also revealed the presence of diverse regulatory elements for the transcriptional and translational control of individual TU, such as promoters, 5¢-UTRs, terminators, 3¢-UTRs, and riboswitches. The conserved promoter sequences for transcription initiation were identified from 2,361 transcription start sites as 5¢-TANNNT and 5¢-TGAC for -10 and -35 elements, respectively. Interestingly, the -35 element and spacer length between them were critical for transcriptional regulation of functionally distinct genes. Total 2,017 transcription termination sites were detected from Term-Seq analysis, revealing that stem structure formation is a prerequisite for transcription termination and that Rho-independent termination prevails in S. avermitilis. Lastly, the TU architecture suggests the presence of novel small RNAs and cis-regulatory elements in the genome. Our findings will serve as invaluable resources for comprehensive understanding on regulatory features of S. avermitilis. Moreover, it is anticipated the elevation of its potential as the heterologous expression host for diverse secondary metabolite biosynthesis.
Project description:The present work aimed at selecting appropriate native temporal promoters with the identical profile of the cumate-inducible promoter under the optimal induction condition. This strategy enabled us to optimized the expression of biosynthetic gene cluster of desired secondary metabolites in Streptomyces, while avoid the use of inducer during fermentation. By clustering the genes with quite similar transcriptional profile with that of gfp controlled by the cumate-inducible promoter based on time-series microarray data, 50 qualified genes were identified, which were controlled by 24 putative promoters. Then, the cumate-inducible promoter were replaced by the 24 native temporal promoters, and fermentation results showed the good performance of these promoters compared to that of the cumate-inducible promoter. Therefore, our strategy could be used to fine-tune the expression of target BGCs for production improvement.
