Abstract
BackgroundThe gram-positive bacterium, Streptomyces avermitilis, holds industrial importance as the producer of avermectin, a widely used anthelmintic agent, and a heterologous expression host of secondary metabolite-biosynthetic gene clusters. Despite its industrial importance, S. avermitilis’ genome organization and regulation of gene expression remain poorly understood. In this study, four different types of Next-Generation Sequencing techniques, including dRNA-Seq, Term-Seq, RNA-Seq and ribosome profiling, were applied to S. avermitilis to determine transcription units of S. avermitilis at a genome-wide level and elucidate regulatory elements for transcriptional and translational control of individual transcription units.ResultBy applying dRNA-Seq and Term-Seq to S. avermitilis MA-4680, a total of 2361 transcription start sites and 2017 transcript 3′-end positions were identified, respectively, leading to determination of 1601 transcription units encoded in S. avermitilis’ genome. Cataloguing the transcription units and integrated analysis of multiple high-throughput data types revealed the presence of diverse regulatory elements for gene expression, such as promoters, 5′-UTRs, terminators, 3′-UTRs and riboswitches. The conserved promoter motifs were identified from 2361 transcription start sites as 5′-TANNNT and 5′-BTGACN for the − 10 and − 35 elements, respectively. The − 35 element and spacer lengths between − 10 and − 35 elements were critical for transcriptional regulation of functionally distinct genes, suggesting the involvement of unique sigma factors. In addition, regulatory sequences recognized by antibiotic regulatory proteins were identified from the transcription start site information. Analysis of the 3′-end of RNA transcript revealed that stem structure formation is a major determinant for transcription termination of most transcription units.ConclusionsThe transcription unit architecture elucidated from the transcripts’ boundary information provides insights for unique genetic regulatory mechanisms of S. avermitilis. Our findings will elevate S. avermitilis’ potential as a production host for a diverse set of secondary metabolites.
Highlights
The gram-positive bacterium, Streptomyces avermitilis, holds industrial importance as the producer of avermectin, a widely used anthelmintic agent, and a heterologous expression host of secondary metabolite-bio‐ synthetic gene clusters
We provide fundamental information on the genome-wide transcription unit (TU) architecture of S. avermitilis determined from transcription start site (TSS) and transcript 3′-end position (TEP) information acquired by differential RNA-Seq and TermSeq, respectively. dRNA-Seq reveals the 5′-end positions of transcripts and differentiate the TSSs from processed 5′-ends by identifying the presence of 5′-triphosphate, which is a typical characteristic of bacterial primary transcripts
Genome‐wide identification of transcription start sites By exploiting dRNA-Seq, we experimentally identified TSSs in the S. avermitilis genome
Summary
The gram-positive bacterium, Streptomyces avermitilis, holds industrial importance as the producer of avermectin, a widely used anthelmintic agent, and a heterologous expression host of secondary metabolite-bio‐ synthetic gene clusters. S. avermitilis can serve as a versatile host for the heterologous production of secondary metabolites from other Streptomyces species [5] Such heterologous expression improves the production yield of useful secondary metabolites [6] and enables the production of novel bioactive derivatives of existing secondary metabolites from reconstructed biosynthetic gene clusters (BGCs) [7]. As a producer and heterologous expression host of important anthelmintics and other secondary metabolites, S. avermitilis is worth to be investigated to increase its potential for secondary metabolite production and activate cryptic BGCs. diverse regulatory mechanisms of secondary metabolism have been reported, most of these are confined to characterized compounds with proven value [14,15,16]. The interpretation of diverse regulatory elements governing gene expression is required
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