Mannose- and Mannobiose-Specific Responses of the Insect-Associated Cellulolytic Bacterium Streptomyces sp. Strain SirexAA-E

Abstract

The cellulolytic insect symbiont bacterium, Streptomyces sp. SirexAA-E (SirexAA-E) secretes a suite of Carbohydrate Active enZymes (CAZymes), which are involved in the degradation of various polysaccharides in the plant cell wall, in response to the available carbon sources. Here, we examined a poorly understood response of this bacterium to mannan, one of the major plant cell wall components. SirexAA-E grew well on mannose, carboxymethyl cellulose (CMC), and locust bean gum (LBG) as sole carbon sources in the culture medium. The secreted proteins from each culture supernatant were tested for their polysaccharide-degrading ability, and the composition of secreted CAZymes in each sample was determined by LC-MS/MS. The results indicated that mannose, LBG, and CMC induced the secretion of mannan and cellulose-degrading enzymes. Interestingly, two α-1,2-mannosidases were abundantly secreted during growth on mannose and LBG. By genomic analysis, we found a unique 12 bp palindromic sequence motif at 4 locations in the SirexAA-E genome, two of which were found upstream of the above-mentioned α-1,2-mannosidase genes, along with a newly identified mannose and mannobiose-responsive transcriptional regulator, SsManR. Furthermore, the previously reported cellobiose-responsive repressor, SsCebR, was determined to also use mannobiose as an effector ligand. To test whether mannobiose induces the sets of genes under the control of the two regulators, SirexAA-E was grown on mannobiose, and the secretome composition was analyzed. As hypothesized, the composition of the mannobiose secretome combined sets of CAZymes found in both LBG and CMC secretomes, and so are likely under the regulation of both SsManR and SsCebR. Importance Streptomyces sp. SirexAA-E, a microbial symbiont of biomass harvesting insects, secretes a suite of polysaccharide-degrading enzymes dependent on the available carbon sources. However, the response of this bacterium to mannan has not been documented. In this study, we investigated the response of this bacterium to mannose, mannobiose, and galactomannan (LBG). By combining biochemical, proteomic, and genomic approaches, we discovered a novel mannose and mannobiose responsive transcriptional regulator, SsManR, which selectively regulates three α-1,2-mannosidase-coding genes. We also demonstrated that the previously described cellobiose responsive regulator, SsCebR, could use mannobiose as an effector ligand. Overall, our findings suggest that the Streptomyces sp. SirexAA-E responds to mannose and mannooligosaccharides through two different transcriptional repressors that regulate the secretion of the plant cell wall-degrading enzymes to extract carbon sources in the host environment.

Effects of mulching film on soil microbial diversity and community of cotton

“…In particular, the abundance of Streptomyces was higher in the DPBOS than PEBOS treatment in non-rhizosphere soil. Streptomyces is able to degrade cellulose, pesticides, and petroleum compounds (Chen et al 2020 ; Ohashi et al 2021 ; Santillan et al 2020 ). Further, Streptomyces may be critically involved in plastics’ degradation.…”

Removal of primary nutrient degraders reduces growth of soil microbial communities with genomic redundancy

“…Supporting Streptomyces as a sharing species our analysis identified mannobiose as a metabolite that was much more abundant when Streptomyces was present vs. in a community lacking Streptomyces . This C source has been shown to be produced by Streptomyces and consumed by Rhodococcus ( Ohashi et al, 2021 ).…”

AURTHO: autoregulation as facilitator of cis-acting element discovery of orthologous transcription factors

“…In total, in 182 Streptomyces strains, 4403 LacI TFs were identified, grouped into 167 COGs. Among these, only 5 (~3% of all LacI TFs) have been subject to studies in Streptomyces species, i.e., i) the galactomannan/mannobiose/mannose utilization repressor ManR (LacI003 in Table 1, conserved in 177/182 species) (Ohashi et al, 2021), (ii) the maltose/maltodextrin catabolism pathway regulator MalR (LacI005 in Table 1, conserved in 176/182 species) (Nguyen, 1999;Nguyen et al, 1997;Schlösser et al, 2001;van Wezel, White, Bibb, et al, 1997;van Wezel, White, Young, et al, 1997), iii) the cellulose/cello-oligosaccharide utilisation regulator CebR (LacI006 in Table 1, conserved in 153/182 species) (Book et al, 2016;Francis et al, 2015;Jourdan et al, 2016;Marushima et al, 2009;Schlösser et al, 2000), iv) the xylan/xylo-oligosaccharide utilization repressor BxlR (LacI015 in Table 1, conserved in 88/182 species) (Giannotta et al, 1996(Giannotta et al, , 2003Tsujibo et al, 2004), and v) the agar-utilisation regulator DagR (LacI139 in Table 1) (Tsevelkhoroloo et al, 2021), the latter being one of the rarest LacI TF, only conserved in two Streptomyces species. Strikingly, the function of the two most conserved LacI TFs (LacI001 and LacI002 in Table 1) is unknown, further illustrating the lack of knowledge about transcriptional regulation in this well-studied bacterial genus.…”

“…The copyright holder for this preprint this version posted April 10, 2022. ; https://doi.org/10.1101/2022.04.06.487287 doi: bioRxiv preprint to their experimentally identified consensus sequences, i.e., GACAACGTTGTC (Ohashi et al, 2021), TGGGAGCGCTCCCA (Schlösser et al, 2000), and CGAA-Nx-TTCG (Giannotta et al, 1996(Giannotta et al, , 2003Tsujibo et al, 2004), respectively. For MalR (LacI005), the two binding sites deduced by DNase footprinting assays (Schlösser et al, 2001) were also found (see Table 1), further confirming that our approach is appropriate for deducing over-represented motifs that closely relate to the ones that were experimentally identified.…”

“…DNA-protein interactions were observed using an ImageQuant TM LAS 4000, by detecting the fluorescence emission of the Cy5-tag using a 670 nm detection filter. ManR (LacI 003, Figure 3 second panel) was used as a positive control for the EMSA method, as its cognate palindromic motif GACAACGTTGTC has been recently confirmed experimentally (Ohashi et al, 2021).…”