A relatively small number of species in the large genus Streptomyces are pathogenic; the best characterized of these is Streptomyces scabies. The pathogenicity of S. scabies strains is dependent on the production of the nitrated diketopiperazine thaxtomin A, which is a potent plant cellulose synthesis inhibitor. Much is known about the genetic loci associated with plant virulence; however, the molecular mechanisms by which S. scabies triggers expression of thaxtomin biosynthetic genes, beyond the pathway-specific activator TxtR, are not well understood. In this study, we demonstrate that binding sites for the cellulose utilization repressor CebR occur and function within the thaxtomin biosynthetic cluster. This was an unexpected result, as CebR is devoted to primary metabolism and nutritive functions in nonpathogenic streptomycetes. In S. scabies, cellobiose and cellotriose inhibit the DNA-binding ability of CebR, leading to an increased expression of the thaxtomin biosynthetic and regulatory genes txtA, txtB, and txtR. Deletion of cebR results in constitutive thaxtomin A production and hypervirulence of S. scabies. The pathogenicity of S. scabies is thus under dual direct positive and negative transcriptional control where CebR is the cellobiose-sensing key that locks the expression of txtR, the key necessary to unlock the production of the phytotoxin. Interestingly, CebR-binding sites also lie upstream of and within the thaxtomin biosynthetic clusters in Streptomyces turgidiscabies and Streptomyces acidiscabies, suggesting that CebR is most likely an important regulator of virulence in these plant-pathogenic species as well.

Streptomyces scabies is an economically important plant pathogen well-known for damaging root and tuber crops by causing scab lesions. Thaxtomin A is the main causative agent responsible for the pathogenicity of S. scabies and cello-oligosaccharides are environmental triggers that induce the production of this phytotoxin. How cello-oligosaccharides are sensed or transported in order to induce the virulent behavior of S. scabies? Here we report that the cellobiose and cellotriose binding protein CebE, and MsiK, the ATPase providing energy for carbohydrates transport, are the protagonists of the cello-oligosaccharide mediated induction of thaxtomin production in S. scabies. Our work provides the first example where the transport and not the sensing of major constituents of the plant host is the central mechanism associated with virulence of the pathogen. Our results allow to draw a complete pathway from signal transport to phytotoxin production where each step of the cascade is controlled by CebR, the cellulose utilization regulator. We propose the high affinity of CebE to cellotriose as possible adaptation of S. scabies to colonize expanding plant tissue. Our work further highlights how genes associated with primary metabolism in nonpathogenic Streptomyces species have been recruited as basic elements of virulence in plant pathogenic species.

Iron is one of the most abundant elements on earth but is found in poorly soluble forms hardly accessible to microorganisms. To subsist, they have developed iron-chelating molecules called siderophores that capture this element in the environment and the resulting complexes are internalized by specific uptake systems. While biosynthesis of siderophores in many bacteria is regulated by iron availability and oxidative stress, we describe here a new type of regulation of siderophore production. We show that in Streptomyces coelicolor, their production is also controlled by N-acetylglucosamine (GlcNAc) via the direct transcriptional repression of the iron utilization repressor dmdR1 by DasR, the GlcNAc utilization regulator. This regulatory nutrient-metal relationship is conserved among streptomycetes, which indicates that the link between GlcNAc utilization and iron uptake repression, however unsuspected, is the consequence of a successful evolutionary process. We describe here the molecular basis of a novel inhibitory mechanism of siderophore production that is independent of iron availability. We speculate that the regulatory connection between GlcNAc and siderophores might be associated with the competition for iron between streptomycetes and their fungal soil competitors, whose cell walls are built from the GlcNAc-containing polymer chitin. Alternatively, GlcNAc could emanate from streptomycetes' own peptidoglycan that goes through intense remodelling throughout their life cycle, thereby modulating the iron supply according to specific needs at different stages of their developmental programme.

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2Common scab disease on root and tuber plants is caused by Streptomyces scabies and related 2 3 species which use the cellulose synthase inhibitor thaxtomin A as main phytotoxin. 4Thaxtomin production is primarily triggered by the import of cello-oligosaccharides. Once 2 5 inside the cell, the fate of the cello-oligosaccharides is dichotomized into i) fueling glycolysis 2 6with glucose for the saprophytic lifestyle through the action of β -glucosidase(s) (BG), and ii) reduced production of thaxtomin when the mutant was cultivated on media containing cello- mediated induction of thaxtomin production and suggests that the role of BglC in the route to 4 0 the pathogenic lifestyle of S. scabies is more complex than currently presented.