SummaryThe poles of bacteria exhibit several specialized functions related to the mobilization of DNA and certain proteins. To monitor the infection of Escherichia coli cells by light microscopy, we developed procedures for the tagging of mature bacteriophages with quantum dots. Surprisingly, most of the infecting phages were found attached to the bacterial poles. This was true for a number of temperate and virulent phages of E. coli that use widely different receptors and for phages infecting Yersinia pseudotuberculosis and Vibrio cholerae. The infecting phages colocalized with the polar protein marker IcsA-GFP. ManY, an E. coli protein that is required for phage l DNA injection, was found to localize to the bacterial poles as well. Furthermore, labelling of l DNA during infection revealed that it is injected and replicated at the polar region of infection. The evolutionary benefits that lead to this remarkable preference for polar infections may be related to l's developmental decision as well as to the function of poles in the ability of bacterial cells to communicate with their environment and in gene regulation.
Sinorhizobium meliloti is a gram-negative soil bacterium capable of forming a symbiotic nitrogen-fixing relationship with its plant host, Medicago sativa. Various bacterially produced factors are essential for successful nodulation. For example, at least one of two exopolysaccharides produced by S. meliloti (succinoglycan or EPS II) is required for nodule invasion. Both of these polymers are produced in high-and low-molecular-weight (HMW and LMW, respectively) fractions; however, only the LMW forms of either succinoglycan or EPS II are active in nodule invasion. The production of LMW succinoglycan can be generated by direct synthesis or through the depolymerization of HMW products by the action of two specific endoglycanases, ExsH and ExoK. Here, we show that the ExpR/Sin quorum-sensing system in S. meliloti is involved in the regulation of genes responsible for succinoglycan biosynthesis as well as in the production of LMW succinoglycan. Therefore, quorum sensing, which has been shown to regulate the production of EPS II, also plays an important role in succinoglycan biosynthesis.
In bacteria, cysteines of cytoplasmic proteins, including the essential enzyme ribonucleotide reductase (RNR), are maintained in the reduced state by the thioredoxin and glutathione/glutaredoxin pathways. An Escherichia coli mutant lacking both glutathione reductase and thioredoxin reductase cannot grow because RNR is disulfide bonded and nonfunctional. Here we report that suppressor mutations in the lpdA gene, which encodes the oxidative enzyme lipoamide dehydrogenase required for tricarboxylic acid (TCA) cycle functioning, restore growth to this redox-defective mutant. The suppressor mutations reduce LpdA activity, causing the accumulation of dihydrolipoamide, the reduced protein-bound form of lipoic acid. Dihydrolipoamide can then provide electrons for the reactivation of RNR through reduction of glutaredoxins. Dihydrolipoamide is oxidized in the process, restoring function to the TCA cycle. Thus, two electron transfer pathways are rewired to meet both oxidative and reductive needs of the cell: dihydrolipoamide functionally replaces glutathione, and the glutaredoxins replace LpdA. Both lipoic acid and glutaredoxins act in the reverse manner from their normal cellular functions. Bioinformatic analysis suggests that such activities may also function in other bacteria.disulfide bond | pyruvate dehydrogenase | α-ketoglutarate dehydrogenase | bacterial genomes R eduction-oxidation reactions play key roles in many essential metabolic pathways. Some redox reactions involve the oxidation or reduction of thiol residues either in an enzyme's cysteines or in small redox-active molecules. In Escherichia coli, the thiol-disulfide biology of the cell is compartmentalized; the majority of protein thiols in the periplasm are oxidized (disulfide bonded), and the majority of protein thiols in the cytoplasm are reduced. However, for cytoplasmic enzymes that use cysteines in catalysis of reductive reactions, disulfide bonds do form, albeit transiently. These bonds are rapidly reduced, restoring the enzyme's activity.Two pathways maintain protein thiols in the reduced state in the cytoplasm of E. coli (1). In the thioredoxin pathway, thioredoxin 1 (encoded by trxA) and thioredoxin 2 (trxC) reduce oxidized substrates. The resulting oxidized thioredoxins are then reduced by thioredoxin reductase (encoded by trxB) to regenerate their activity. In the glutathione/glutaredoxin pathway, glutaredoxins 1 (grxA), 2 (grxB), and 3 (grxC) can reduce oxidized substrate proteins. The small molecule thiol glutathione (GSH) and glutathione reductase (gor) provide electrons to maintain glutaredoxins in the reduced state.The importance of the TrxB and Gor pathways is indicated by the finding that when null mutations in certain genes of both pathways are combined, E. coli cannot grow. The reason for this synthetic lethality is that the essential enzyme ribonucleotide reductase (RNR) must be reduced by these pathways to maintain its activity. In certain synthetic lethal combinations of mutations (e.g., gor trxB), but not others (trxA trxC grxA), cel...
The major oxidative stress response in Streptomyces is controlled by the sigma factor SigR and its cognate antisigma factor RsrA, and SigR activity is tightly controlled through multiple mechanisms at both the transcriptional and posttranslational levels. Here we show that sigR has a highly unusual GTC start codon and that this leads to another level of SigR regulation, in which SigR translation is repressed by translation initiation factor 3 (IF3). Changing the GTC to a canonical start codon causes SigR to be overproduced relative to RsrA, resulting in unregulated and constitutive expression of the SigR regulon. Similarly, introducing IF3* mutations that impair its ability to repress SigR translation has the same effect. Thus, the noncanonical GTC sigR start codon and its repression by IF3 are critical for the correct and proper functioning of the oxidative stress regulatory system. sigR and rsrA are cotranscribed and translationally coupled, and it had therefore been assumed that SigR and RsrA are produced in stoichiometric amounts. Here we show that RsrA can be transcribed and translated independently of SigR, present evidence that RsrA is normally produced in excess of SigR, and describe the factors that determine SigR-RsrA stoichiometry.
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