SUCCINOGLYCAN PRODUCTION BY RHIZOBIUM-MELILOTI IS REGULATED THROUGH THE EXOS-CHVI 2-COMPONENT REGULATORY SYSTEM

The Rhizobium meliloti exoS gene is involved in regulating the product ion of succinoglycan, which plays a crucial role in the establishment of the symbiosis between R. meliloti Rm1021 and its host pint, alfalfa . The exoS96::Tn5 mutation causes tile upregulation of the succinoglyc an biosynthetic genes, thereby resulting in the overproduction of succ inoglycan. Through cloning and sequencing, we found that the exoS gene is a close homolog of the Agrobacterium tumefaciens chvG gene, which has been proposed to encode the sensor protein of the ChvG-ChvI two-co mponent regulatory system, a member of the EnvZ-OmpR family. Further a nalyses revealed the existence of a newly discovered A. tumefaciens ch vI homolog located just upstream of the R. meliloti exoS gene, R. meli loti ChvI may serve as the response regulator of ExoS in a two-compone nt regulatory system, By using ExoS-specific antibodies, it was found that the ExoS protein cofractionated with membrane proteins, suggestin g that it is located in the cytoplasmic membrane. By using the same an tibodies, it was shown that the exoS96::Tn5 allele encodes an N-termin al truncated derivative of ExoS. The cytoplasmic histidine kinase doma in of ExoS was expressed in Escherichia coli and purified, as was the R. meliloti CHvI protein, The ChvI protein autophosphorylated inn the presence of acetylphosphate, and the ExoS cytoplasmic domain fragment autophosphorylated at a histidine residue in the presence of ATP. The ChvI protein was phosphorylated in the presence of ATP only when the h istidine kinase domain of ExoS was also present. We propose a model fo r regulation of succinoglycan production by R. meliloti through the Ex oS-ChvI two-component regulatory system.