Expression of the Escherichia coli NRZ nitrate reductase is highly growth phase dependent and is controlled by RpoS, the alternative vegetative sigma factor

Abstract

SummaryIn the absence of oxygen, many bacteria preferentially use nitrate as a terminal electron acceptor for anaerobic respiration. In Escherichia coli, there are two membrane-bound, differentially regulated nitrate reductases. While the physiological basis for this metabolic redundancy is not completely understood, during exponential growth, synthesis of NRA is greatly induced by anaerobiosis plus nitrate, whereas NRZ is expressed at a low level that is not in¯uenced by anaerobiosis or nitrate. In the course of identifying genes controlled by the stationary phase regulatory factor RpoS (s s ),we found that the expression of NRZ is induced during entry into stationary phase and highly dependent on this alternative sigma factor. Expression studies, using operon fusions and nitrate reductase assays, revealed that the NRZ operon is controlled mainly at the level of transcription and is induced 10-fold at the onset of stationary phase in rich media. Consistent with previous reports of RpoS expression, the RpoS dependency of NRZ in minimal media was very high (several hundredfold). We also observed a ®ve-fold stationary phase induction of NRZ in an rpoS background, indicating that other regulatory factors, besides RpoS, are probably involved in transcriptional control of NRZ. The RpoS dependence of NRZ expression was con®rmed by Northern analyses using RNA extracted from wild-type and rpoS À strains sampled in exponential and stationary phase. In toto, these data indicate that RpoS-mediated regulation of NRZ may be an important physiological adaptation that allows the cell to use nitrate under stress-associated conditions.

Periplasmic Nitrate Reductase (NapABC Enzyme) Supports Anaerobic Respiration byEscherichia coliK-12

“…The NarZYV enzyme contributed little to the overall nitrate reductase activity (compare the activities of strains VJS5435 and VJS5437), as expected since low levels of this enzyme are synthesized during exponential growth (10). The NarG ϩ NapA ϩ strain VJS5432 exhibited significant endogenous activity, which increased fourto fivefold when reduced viologen was provided as an artificial electron donor.…”

Role of the Escherichia coli nitrate transport protein, NarU, in survival during severe nutrient starvation and slow growth

“…In contrast, the third, nitrate reductase Z, is expressed during exponential growth at such a low level that it contributes insignificantly to the total rate of nitrate reduction when either of the other two enzymes is also expressed. However, in both E. coli and S. typhimurium, its synthesis is induced under the control of RpoS during the stationary phase of growth (Chang et al, 1999;Spector et al, 1999). In this study, we demonstrated that NarU also accumulates during the stationary phase rather than during exponential growth, that its synthesis confers a selective advantage during nutrient starvation or very slow growth, and that narU is cotranscribed with narZ, implicating narU as the first gene of a five-gene polycistronic narUZYWV operon.…”

“…The third nitrate reductase, encoded by the narZYWV operon, is structurally very similar to nitrate reductase A Bonnefoy et al, 1997), but is expressed extremely weakly during both aerobic and anaerobic growth (Iobbi et al, 1987). Although expression of nitrate reductase Z is induced during entry into the stationary phase of growth, its activity is still at the lower limits of detection by most biochemical assays (Iobbi-Nivol et al, 1990;Bonnefoy et al, 1997;Chang et al, 1999;. Consequently, nitrate reductase Z contributes very little to the overall rate of nitrate reduction by E. coli, and is sufficient to support only a very low rate of nitrate-dependent anaerobic growth with glycerol as the non-fermentable carbon source .…”

RpoS-Regulated Genes ofEscherichia coliIdentified by RandomlacZFusion Mutagenesis

“…This was found to be important, as many of the genes identified have paralogous functions whose activity may be masked by the presence of another gene product (e.g., NarG or NarZ). There are several published RpoS-dependent paralogs, including NarZ (nitrate reductase) (18) and Ldc (lysine decarboxylase) (89), that comprise only a small percentage of the cell's total activity. There are many other paralogous functions that are controlled by RpoS, and collectively this suggests that expressing a subset of key duplicated metabolic functions is an important physiological imperative in stationary-phase cultures.…”

“…Nitrate reductase Z is encoded by members of the nar-ZYWV operon (71). The RpoS-dependent narY gene codes for the beta subunit of nitrate reductase Z, which functions as an electron acceptor during anaerobic growth on nitrate in E. coli (18).…”