RESUMO
Osmoregulated periplasmic glucans (OPGs) are a family of periplasmic oligosaccharides found in the envelope of most Proteobacteria. They are required for virulence of zoo- and phyto-pathogens. The glucose backbone of OPGs is substituted by various kinds of molecules depending on the species, O-succinyl residues being the most widely distributed. In our model, Dickeya dadantii, a phytopathogenic bacteria causing soft rot disease in a wide range of plant species, the backbone of OPGs is substituted by O-succinyl residues in media of high osmolarity and by O-acetyl residues whatever the osmolarity. The opgC gene encoding a transmembrane protein required for the succinylation of the OPGs in D. dadantii was found after an in silico search of a gene encoding a protein with the main characteristics recovered in the two previously characterized OpgC of E. coli and R. sphaeroides, i.e. 10 transmembrane segments and one acyl-transferase domain. Characterization of the opgC gene revealed that high osmolarity expression of the succinyl transferase is controlled by both the EnvZ-OmpR and RcsCDB phosphorelay systems. The loss of O-succinyl residue did not affect the virulence of D. dadantii, suggesting that only the glucose backbone of OPGs is required for virulence.
Assuntos
Proteínas de Bactérias/genética , Enterobacteriaceae/genética , Enterobacteriaceae/metabolismo , Glucanos/metabolismo , Osmorregulação , Periplasma/metabolismo , Proteínas de Bactérias/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Ordem dos Genes , Teste de Complementação Genética , Genoma Bacteriano , Concentração Osmolar , Fosforilação , Regiões Promotoras Genéticas , Ligação Proteica , Virulência/genéticaRESUMO
The opgGH operon encodes glucosyltransferases that synthesize osmoregulated periplasmic glucans (OPGs) from UDP-glucose, using acyl carrier protein (ACP) as a cofactor. OPGs are required for motility, biofilm formation, and virulence in various bacteria. OpgH also sequesters FtsZ in order to regulate cell size according to nutrient availability. Yersinia pestis (the agent of flea-borne plague) lost the opgGH operon during its emergence from the enteropathogen Yersinia pseudotuberculosis. When expressed in OPG-negative strains of Escherichia coli and Dickeya dadantii, opgGH from Y. pseudotuberculosis restored OPGs synthesis, motility, and virulence. However, Y. pseudotuberculosis did not produce OPGs (i) under various growth conditions or (ii) when overexpressing its opgGH operon, its galUF operon (governing UDP-glucose), or the opgGH operon or Acp from E. coli. A ΔopgGH Y. pseudotuberculosis strain showed normal motility, biofilm formation, resistance to polymyxin and macrophages, and virulence but was smaller. Consistently, Y. pestis was smaller than Y. pseudotuberculosis when cultured at ≥ 37°C, except when the plague bacillus expressed opgGH. Y. pestis expressing opgGH grew normally in serum and within macrophages and was fully virulent in mice, suggesting that small cell size was not advantageous in the mammalian host. Lastly, Y. pestis expressing opgGH was able to infect Xenopsylla cheopis fleas normally. Our results suggest an evolutionary scenario whereby an ancestral Yersinia strain lost a factor required for OPG biosynthesis but kept opgGH (to regulate cell size). The opgGH operon was presumably then lost because OpgH-dependent cell size control became unnecessary.
Assuntos
Proteínas de Bactérias/genética , Evolução Molecular , Yersinia pestis/patogenicidade , Yersinia pseudotuberculosis/genética , Yersinia pseudotuberculosis/patogenicidade , Animais , Western Blotting , Modelos Animais de Doenças , Deleção de Genes , Glucanos/biossíntese , Glucanos/genética , Camundongos , Óperon/genética , Proteínas Periplásmicas/biossíntese , Proteínas Periplásmicas/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Osmoregulated periplasmic glucans (OPGs) are oligosaccharides found in the periplasm of many Gram-negative bacteria. Glucose is the sole constitutive sugar and this backbone may be substituted by various kinds of molecules depending on the species. In E. coli, OPG are substituted by phosphoglycerol and phosphoethanolamine derived from membrane phospholipids and by succinyl residues. In this study, we describe the isolation of the opgE gene encoding the phosphoethanolamine transferase by a screen previously used for the isolation of the opgB gene encoding the phosphoglycerol transferase. Both genes show structural and functional similarities without sequence similarity.
Assuntos
Etanolaminofosfotransferase/genética , Glucanos/biossíntese , Glucose/metabolismo , Transferases (Outros Grupos de Fosfato Substituídos)/genética , Escherichia coli/genética , Etanolaminofosfotransferase/química , Etanolaminofosfotransferase/isolamento & purificação , Etanolaminas/metabolismo , Regulação Bacteriana da Expressão Gênica , Glucanos/genética , Glucose/genética , Periplasma/genética , Periplasma/metabolismo , Conformação Proteica , Homologia Estrutural de Proteína , Transferases (Outros Grupos de Fosfato Substituídos)/químicaRESUMO
Osmoregulated periplasmic glucans (OPGs) are general constituents of many Proteobacteria. Synthesis of these oligosaccharides is repressed by increased osmolarity of the medium. OPGs are important factors required for full virulence in many zoo- or phytopathogens including Dickeya dadantii. The phytopathogen enterobacterium D. dadantii causes soft-rot disease on a wide range of plant species. The total loss of virulence of opg-negative strains of D. dadantii is linked to the constitutive activation of the RcsCD RcsB phosphorelay highlighting relationship between this phosphorelay and OPGs. Here we show that OPGs control the RcsCD RcsB activation in a concentration-dependent manner, are required for proper activation of this phosphorelay by medium osmolarity, and a high concentration of OPGs in planta is maintained to achieve the low level of activation of the RcsCD RcsB phosphorelay required for full virulence in D. dadantii.