Gene expression in Sinorhizobium meliloti during nutrient deprivation

To persist, bacteria must adapt dynamically to their environment by switching on or off different suites of genes in response to their surroundings. Two major parameters that bacteria constantly monitor are nutrient status and oxygen tension. In soil, these resources are often scarce. Nutrients enter this ecosystem intermittently, however the diverse bacterial populations compete for these nutrients and they are quickly utilized. Consequently, the natural physiological state of indigenous soil bacteria is either dormancy or negligible growth. Understanding how bacteria are able to monitor and respond to their environment in this nutrient-deprived state is fundamental to our understanding of microbial biology. By mutagenizing the genome of Sinorhizobium meliloti with a Tn5 derivative (Tn5luxAB), which generates transcriptional fusions resulting in bacterial bioluminescence when the gene fusion is expressed, a gene was identified in S. meliloti that is induced by environmental parameters which are representative of life in soil; that is, nitrogen or carbon deprivation, low oxygen tension, as well as during post-exponential stationary-phase growth, and by osmotic stress. The tagged gene was found to be part of an operon consisting of two open reading frames (ORF), which were designated ndiA and ndiB for nutrient deprivation induced genes. Comparison of the deduced amino acid sequences of both ndiA and ndiB to the protein databases did not reveal similarities with any known genes; therefore, they appear to be novel. In addition, a gene involved in the regulation of this operon was isolated, by carrying out a second round of mutagenesis on the primary mutant strain C22 with a Tn3 derivative, Tn1721. A library of 3000 double mutant derivatives of C22 was screened for strains with altered luciferase expression patterns. One double mutant that failed to express the C22 luciferase fusion under any of the conditions tested was identified. This mutant contained a Tn1721 insertion in a gene which encodes a protein with a high degree of similarity to the tryptophan-rich sensory protein, TspO, from Rhodobacter sphaeroides , as well as to the mitochondrial benzodiazepine receptor, pK18. Furthermore, proper environmental control of the ndi-luxAB reporter gene fusion was found to be restored after introduction of the tspO coding region in trans, under all inducing conditions tested. Thus, the experiments described here showed both the presence of a novel operon whose expression is induced by multiple environmental (stress) conditions, as well as a hitherto unidentified S. meliloti sensor/regulator locus involved in environmental control of gene expression.