System-Level Analysis of Transcriptional and Translational Regulatory Elements in Streptomyces griseus

Abstract

Bacteria belonging to Streptomyces have the ability to produce a wide range of secondary metabolites through a shift from primary to secondary metabolism regulated by complex networks activated after vegetative growth terminates. Despite considerable effort to understand the regulatory elements governing gene expression related to primary and secondary metabolism in Streptomyces, system-level information remains limited. In this study, we integrated four multi-omics datasets from Streptomyces griseus NBRC 13350: RNA-seq, ribosome profiling, dRNA-seq, and Term-Seq, to analyze the regulatory elements of transcription and translation of differentially expressed genes during cell growth. With the functional enrichment of gene expression in different growth phases, one sigma factor regulon and four transcription factor regulons governing differential gene transcription patterns were found. In addition, the regulatory elements of transcription termination and post-transcriptional processing at transcript 3′-end positions were elucidated, including their conserved motifs, stem-loop RNA structures, and non-terminal locations within the polycistronic operons, and the potential regulatory elements of translation initiation and elongation such as 5′-UTR length, RNA structures at ribosome-bound sites, and codon usage were investigated. This comprehensive genetic information provides a foundational genetic resource for strain engineering to enhance secondary metabolite production in Streptomyces.

Characterization of bacterial intrinsic transcription terminators identified with TERMITe – a novel method for comprehensive analysis of Term-seq data

“…E. coli was observed to have much higher enrichment, while the highest was observed for the rest of the analyzed bacterial species. The rarity of the strong U-rich tracts in Streptomyces genomes was noted previously (18–22). It has also been reported that terminators with U-rich tracts tend to terminate more strongly in S. avermitilis (18).…”

“…In our analysis we included 691 and 957 intrinsic termination sites predicted by TERMITe in E. coli based on the data described in this study and Choe et al (17), respectively; 635, 1165, 984 and 1214 sites of B. subtilis based on data from Chhabra et al (15), Mandell et al (7), Dar et al (14) and Mondal et al (8), respectively; 796 terminators from E. faecalis (14); 862 terminators from L. monocytogenes (14); 206 terminators from Z. mobilis (24); 165 terminators from Synechocystis sp . (16); 629 terminators from S. coelicolor (20), 573 terminators from S. venezuelae (20); 520 terminators from S. tsukubensiis (20); 737 terminators from S. griseus (22); 495 terminators from S. lividans (20); 435 terminators from S. clavuligerus (21); 707 terminators from S. avermitilis (20) (Supplementary Figure 3A). All of the above-listed terminators have been identified using TERMITe as described in the Methods section.…”

“…follow a bimodal distribution. It was previously suggested that such distribution is related to the evolution of two distinct mechanisms of transcription termination (Rho-dependent and Rho-independent/intrinsic) (16, 21, 22). Thus, the observed bimodality might suggest that the Rho-dependent mechanism might be utilized more often in the GC-rich Streptomyces genus than in other bacteria, having less GC-rich genomes, which is in agreement with the observed lowest fraction of intrinsic terminators among all identified 3’ RNA termini.…”

“…Next, we used TERMITe to perform a comprehensive analysis of the reported here and publicly available Term-seq datasets to investigate the variability in intrinsic termination signals across bacterial species. In our analysis we included 691 and 957 intrinsic termination sites predicted by TERMITe in E. coli based on the data described in this study and Choe et al (17) (20), 573 terminators from S. venezuelae (20); 520 terminators from S. tsukubensiis (20); 737 terminators from S. griseus (22); 495 terminators from S. lividans (20); 435 terminators from S. clavuligerus (21); 707 terminators from S. avermitilis (20) (Supplementary Figure 3A). All of the above-listed terminators have been identified using TERMITe as described in the Methods section.…”

“…(16), E. coli (17), Streptomyces sp. (18)(19)(20)(21)(22), Pseudomonas aeruginosa (23), Zymomonas mobilis (24), Methanosarcina mazei, and Sulfolobus acidocaldarius (25). Most studies rely on in-house custom analytical approaches to identify and analyze either RNA decay products or transcription termination sites.…”