The chitinolytic cascade in Vibrios is regulated by chitin oligosaccharides and a two-component chitin catabolic sensor/kinase

Abstract

Chitin, a highly insoluble polymer of GlcNAc, is produced in massive quantities in the marine environment. Fortunately for survival of aquatic ecosystems, chitin is rapidly catabolized by marine bacteria. Here we describe a bacterial two-component hybrid sensor͞ kinase (of the ArcB type) that rigorously controls expression of Ϸ50 genes, many involved in chitin degradation. The sensor gene, chiS, was identified in Vibrio furnissii and Vibrio cholerae (predicted amino acid sequences, full-length: 84% identical, 93% similar). Mutants of chiS grew normally on GlcNAc but did not express extracellular chitinase, a specific chitoporin, or ␤-hexosaminidases, nor did they exhibit chemotaxis, transport, or growth on chitin oligosaccharides such as (GlcNAc) 2. Expression of these systems requires three components: wild-type chiS; a periplasmic highaffinity chitin oligosaccharide, (GlcNAc) n (n > 1), binding protein (CBP); and the environmental signal, (GlcNAc) n. Our data are consistent with the following model. In the uninduced state, CBP binds to the periplasmic domain of ChiS and ''locks'' it into the minus conformation. The environmental signal, (GlcNAc) n, dissociates the complex by binding to CBP, releasing ChiS, yielding the plus phenotype (expression of chitinolytic genes). In V. cholerae, a cluster of 10 contiguous genes (VC0620 -VC0611) apparently comprise a (GlcNAc) 2 catabolic operon. CBP is encoded by the first, VC0620, whereas VC0619 -VC0616 encode a (GlcNAc) 2 ABC-type permease. Regulation of chiS requires expression of CBP but not (GlcNAc) 2 transport. (GlcNAc)n is suggested to be essential for signaling these cells that chitin is in the microenvironment.

Global occurrence and heterogeneity of the Roseobacter-clade species Ruegeria mobilis

“…Two of our R. mobilis strains were isolated from copepods, however, chitinases were not detected in the genomes. Instead, we identified the complete uptake system for N-acetylglucosamine dimers including the sensor chiS (Li and Roseman, 2004;Machado et al, 2015). This could present an example of R. mobilis not only being a supplier of nutrients, but also benefiting from the leaky production by others.…”

Identification and Functional Characterization of a Novel OprD-like Chitin Uptake Channel in Non-chitinolytic Bacteria

“…Vibrio species possess competent chitin degradation and uptake systems that allow the bacteria to metabolize chitinous materials, generating catabolic intermediates that can be used as their sole source of energy (5,(41)(42)(43)(44)(45)(46)(47). In marked contrast, E. coli is a non-chitinolytic bacterium living primarily in the gastrointestinal tract of animals, and its generation of cellular energy relies on glucose-enriched nutrients.…”

“…Solvent-free bilayer (Montal-Mueller type) formation was performed using 1,2-diphytanoyl-sn-glycero-3-phosphatidylcholine; Avanti Polar Lipids, Alabaster, AL). First, the aperture was prepainted with a few microliters of 1% (v/v) hexadecane in hexane, then a planar bilayer was formed across the aperture by lowering and raising the liquid level (23 5 ) and chitohexaose (GlcNAc 6 ). Each sugar was added to the cis side of the chamber.…”

Chitoporin from Vibrio harveyi, a Channel with Exceptional Sugar Specificity

“…In marine ecosystems, chitin-containing substances are major sources of carbon and nitrogen for marine vibrios. The catabolic cascade of chitin utilization has been proposed to be a complex system, involving a large number of genes that are orchestrated under the stringent control of the chitin-induced operon (4). In brief, the multistep chitin degradation pathway involves (i) chitin sensing and degradation; (ii) chitooligosaccharide uptake into the periplasm; (iii) degradation of the transported products to GlcNAc and GlcNAc 2 , which are then transported farther into the cytoplasm; and (iv) conversion of GlcNAc intermediates to Fru-6-P and NH 3 ϩ , which are metabolized for energy production and biosynthesis (5)(6)(7).…”

“…However, the specific function of chitoporin as a sugar-specific channel had never been clearly elucidated. We recently employed black lipid membrane (BLM) 4 reconstitution and liposome swelling assays to demonstrate that chitoporin from V. harveyi (VhChiP) is a pore-forming channel that performs highly specific translocation of chitooligosaccharides (11). In the present study, we focused on the kinetic evaluation of single chitooligosaccharide translocation through a single VhChiP.…”