The exuT Gene of Erwinia chrysanthemi EC16: Nucleotide Sequence, Expression, Localization, and Relevance of the Gene Product

Abstract

Galacturonic acid (GalUA) is a major component of pectin and polygalacturonic acid in the plant cell wall. In the phytopathogen Erwinia chrysanthemi, the uptake of molecules derived from degradation of these polymers is an important early step in the events preceding induction of pectinases, ultimately leading to plant tissue maceration. Uptake systems for GalUA and dimers of GalUA have been described and shown to be inducible in E. chrysanthemi. The GalUA uptake gene (exuT) was cloned and sequenced. Nucleotide sequence analysis identified an open reading frame encoding a 345-amino-acid polypeptide with a calculated mass of 37,825 Da. This polypeptide is predicted to be an integral membrane protein based on its high nonpolar amino acid content and hydropathic profile. Localization studies with the labeled polypeptide in the T7-RNA polymerase system also suggest that ExuT is a membrane protein. This evidence is further supported by the observation of hybrid ExuT-PhoA proteins in the bacterial cytoplasmic membrane following immunoblot analysis. Northern (RNA) analysis indicated that the gene is inducible in the presence of the monomer, GalUA. A targeted mutation in the exuT gene affected the utilization of GalUA as a role carbon source for growth. Maceration of potato tuber tissue by this mutant was delayed and reduced, when compared with the parental strain EC16.

Erwinia chrysanthemi tolCIs Involved in Resistance to Antimicrobial Plant Chemicals and Is Essential for Phytopathogenesis†

“…E. chrysanthemi outer membranes were prepared as previously described (23,18). Following protein separation on SDS-10% polyacrylamide gels and transfer to polyvinylidene difluoride (PVDF) membranes, immunological analysis was performed (using E. coli anti-TolC rabbit polyclonal antiserum) on outer-membrane preparations and detected with alkaline phosphatase-conjugated goat anti-rabbit immunoglobulin G and 5-bromo-4-chloro-3-indolyl phosphate-nitroblue tetrazolium (Sigma Co.).…”

“…Production of plant cell wall-degrading enzymes and avirulence proteins by the phytopathogen contributes to the destruction of plant structural barriers and evasion of certain host defense responses, respectively (4, 9, 10, 21, 37). Nutrient acquisition in E. chrysanthemi from degraded plant cell-wall pectin is facilitated by transport systems for pectin-derived oligomers and monomers (18,20,22,46). Secretion of pathogenesis-related proteins across the bacterial cytoplasmic and outer membranes requires several export systems that have been extensively studied.…”

Regulation of D-galacturonate metabolism in <i>Caulobacter crescentus</i> by HumR, a LacI-family transcriptional repressor

“…Within the CC_1487-1495 cluster, a second potential operator sequence upstream of CC_1494-1495 shared 11 out of 14 bp. Other potential operators included a 14/14 bp match located 116 bp upstream of the start codon for CC_1446, the first gene in a three gene unit (CC_1446-1448) annotated as encoding the three key components of an inner membrane TRAP family transport system that could potentially serve as an ATP-independent transporter for hexuronates [27]; a 13/14 bp match located 79 bp upstream of CC_1509, the first gene in a likely operon that includes a homolog of the exuT hexuronate transporter (CC_1508) [28]; an 11/14 bp match located 99 bp upstream of the start codon for CC_3152, which is annotated as encoding a pectatelyase; and an 11/14 bp match located upstream of CC_0442, a putative TonB-dependent outer membrane receptor.…”

Sugar transporter (MfsX) of the major facilitator superfamily is required for flagella-mediated pathogenesis in Dickeya dadantii 3937

“…In the plant pathogenic bacterium D. dadantii 3937, two sugar efflux transporter genes sotA and sotB that belong to the MFS lower the intracellular concentration of toxic sugars or sugar metabolites (Condemine 2000). ExuT, a member of the MFS, was also reported to be involved in the pathogenicity in D. dadantii EC16 (Haseloff et al 1998).…”