The production of the Sinorhizobium meliloti exopolysaccharide, succinoglycan, is required for the formation of infection threads inside root hairs, a critical step during the nodulation of alfalfa (Medicago sativa) by S. meliloti. Two bacterial mutations, exoR95::Tn5 and exoS96::Tn5, resulted in the overproduction of succinoglycan and a reduction in symbiosis. Systematic analyses of the symbiotic phenotypes of the two mutants demonstrated their reduced efficiency of root hair colonization. In addition, both the exoR95 and exoS96 mutations caused a marked reduction in the biosynthesis of flagella and consequent loss of ability of the cells to swarm and swim. Succinoglycan overproduction did not appear to be the cause of the suppression of flagellum biosynthesis. Further analysis indicated that both the exoR95 and exoS96 mutations affected the expression of the flagellum biosynthesis genes. These findings suggest that both the ExoR protein and the ExoS/ChvI two-component regulatory system are involved in the regulation of both succinoglycan and flagellum biosynthesis. These findings provide new avenues of understanding of the physiological changes S. meliloti cells go through during the early stages of symbiosis and of the signal transduction pathways that mediate such changes.Sinorhizobium meliloti and its legume host, alfalfa (Medicago sativa), establish an effective nitrogen-fixing symbiosis through a series of signal exchanges that starts with the exchange of Nod (nodulation) factors and flavonoids, which results in the formation of curled alfalfa root hairs that are colonized by S. meliloti cells (13,20,32). The colonized curled root hairs develop infection threads within the root hairs, which allow S. meliloti cells to invade the developing root nodules (14,16,31). A successful invasion of nodules by S. meliloti will result in the formation of pink nitrogen-fixing nodules. The pink color is due to the presence of leghemoglobin. Nodules that are not occupied by S. meliloti and/or not capable of fixing nitrogen are most often white due to the lack of leghemoglobin (20).The formation of infection threads inside root hairs requires the presence of an S. meliloti exopolysaccharide, succinoglycan (9), in addition to the Nod factor (33). Succinoglycan is a polymer that consists of different numbers of a repeating unit consisting of one galactose and seven glucoses with three modification groups: acetyl, pyruvyl, and succinyl (17, 24). All three modifications must be present in order for the S. meliloti succinoglycan to be active in eliciting infection thread formation (9). Surprisingly, overproduction of succinoglycan appears to reduce efficiency of nodulation (12).Two S. meliloti mutants, exoR95::Tn5 and exoS96::Tn5, were isolated based on their ability to overproduce succinoglycan (12). The exoR gene encodes a protein of 268 amino acids that shares no significant homology with any other protein in currently available databases (23). The exoS gene encodes the membrane-bound sensor of the ExoS/ChvI two-component r...

SUMMARY In Escherichia coli, RNA degradation often begins with conversion of the 5′-terminal triphosphate to a monophosphate, creating a better substrate for internal cleavage by RNase E. Remarkably, no homologue of this key endonuclease is present in many bacterial species, such as Bacillus subtilis and various pathogens. Here we report that the degradation of primary transcripts in B. subtilis can nevertheless be triggered by an analogous process to generate a short-lived, monophosphorylated intermediate. Like its E. coli counterpart, the B. subtilis RNA pyrophosphohydrolase that catalyzes this event is a Nudix protein that prefers unpaired 5′ ends. However, in B. subtilis this modification exposes transcripts to rapid 5′-exonucleolytic degradation by RNase J, which is absent in E. coli but present in most bacteria lacking RNase E. This pathway, which closely resembles the mechanism by which deadenylated mRNA is degraded in eukaryotic cells, explains the stabilizing influence of 5′-terminal stem-loops in such bacteria.

The establishment of an effective nitrogen-fixing symbiosis between Sinorhizobium meliloti and its legume host alfalfa (Medicago sativa) depends on the timely expression of nodulation genes that are controlled by LysR-type regulators. Ninety putative genes coding for LysR-type transcriptional regulators were identified in the recently sequenced S. meliloti genome. All 90 putative lysR genes were mutagenized using plasmid insertions as a first step toward determining their roles in symbiosis. Two new LysR-type symbiosis regulator genes, lsrA and lsrB, were identified in the screening. Both the lsrA and lsrB genes are expressed in free-living S. meliloti cells, but they are not required for cell growth. An lsrA1 mutant was defective in symbiosis and elicited only white nodules that exhibited no nitrogenase activity. Cells of the lsrA1 mutant were recovered from the white nodules, suggesting that the lsrA1 mutant was blocked early in nodulation. An lsrB1 mutant was deficient in symbiosis and elicited a mixture of pink and white nodules on alfalfa plants. These plants exhibited lower overall nitrogenase activity than plants inoculated with the wild-type strain, which is consistent with the fact that most of the alfalfa plants inoculated with the lsrB1 mutant were short and yellow. Cells of the lsrB1 mutant were recovered from both pink and white nodules, suggesting that lsrB1 mutants could be blocked at multiple points during nodulation. The identification of two new LysR-type symbiosis transcriptional regulators provides two new avenues for understanding the complex S. meliloti-alfalfa interactions which occur during symbiosis.