Nitrate enzyme induction

In bacteria, with Pichinoty type A dissimilatory nitrate reductase, induction of the enzyme occurs only under anaerobic conditions, and occasionally anaerobiosis is sufficient to trigger formation in the absence of nitrate (Schulp Md Stouthamer, 1970 Sinclair and White, 1970). However, in these cases of induction by anaerobiosis, the level of enzyme production is greatly enhanced by the presence of nitrate. Dissimilatory nitrate reductase may also be induced under anaerobic conditions by nitrite and azide. 

Section VI indicates that availability of nitrate at the induction and assimilation sites plays a major role in regulating the level of nitrate reductase and the in situ rate of reduction of nitrate. If this is valid, then factors that regulate the uptake, translocation, and entry of nitrate into the cytoplasm of cells of various plant organs have more significant effects on regulating enzyme induction and rate of nitrate assimilation throughout the life cycle of plants than other mechanisms that affect induction, inhibition, and assimilation. 

Judged by enzyme induction, Menary and Jones (1972) indicated that nitrate was unable to move from a storage pool (vacuole) to the cytoplasm of ripe paw paw fruit. Using a modified in vivo assay, Ferrari et al. (1973) estimated sizes of active (metabolic) and inactive (storage) pools of nitrate in tobacco cells, barley aleurone layers and maize leaves. The location and nature of the pools was not defined. 

The NO-ASA family of NO-NSAIDs, notably NCX 4016 [Fig. 20.2(a)] and NCX 4040 [Fig. 20.2(b)], has been extensively researched. These two isomeric hybrid NORMS differ only in the snbstitution of the linker group. Hulsman et al. was the first to comment that in NCX-4040 the presnmed invisible linker is in fact solely responsible for the anti-tumor effect of the molecule and that both the NO-release warhead and the ASA are passive bystanders (Hulsman, Medema et al. 2007). The Unker warhead moiety is efficiently bioactivated to a quinone methide thiophiUc electrophile, in simile with much earUer literature reports (Myers and Widlanski 1993). Molecules in which the nitrate group of NO-ASA was replaced by a comparably good leaving group (termed X-ASA Fig. 20.12) showed very similar properties in vitro with respect to activity (1) cytotoxic/genotoxic (2) antiproliferative (3) chemopreventive (ARE activation - phase 11 enzyme induction) and (4) the anti-inflammatory (Dunlap et al. 2(X)7, 2008). 

It was long believed that bacteria were unique in their ability to denitrify. However, Shoun and Tanimoto (1991) and Shoun et al., (1989) demonstrated that the fungus, Fusarium oxysporum, could be induced to synthesize an enzyme system capable of the anaerobic reduction of nitrite to N2O. Induction occurred under conditions of low oxygen concentrations in the presence of nitrate or nitrite. One and pethaps the only component of this nitrite reductase system is a unique, soluble cytochrome P-450 (P-450dNIR), which is more similar in its cDNA-inferred amino acid sequence to soluble, bacterial P-450 enzymes (espe-... 

Attempts to understand the molecular basis for the induction of synthesis of specific proteins/enzymes are discussed in most of the chapters, but some topics are more advanced in this respect than others. In most cases induction is due to activation of gene expression, as measured by increased steady state levels of mRNA, but in some cases, for instance nitrate reductase, post-transcriptional events may also be important. The eventual aim of course is to understand the series of events - the so-called signal transduction pathway - which lead from perception of the signal to increased mRNA levels and hence increased enzyme synthesis. In a... 

The enzyme activity is induced by both NOj" and Mo when in the presence of each other. The induction of enzyme activity by N03" is a slow process and requires mRNA-dependent synthesis of apoprotein, whereas the induction of enzyme activity by Mo is much faster, as it involves only rapid activation of the apoprotein by Mo (Jones et al., 1976, 1978). Notion and Hewitt (1979) showed that the Mo-free apoenzyme could be activated by addition of Mo complex obtained from acid washings of the native enzyme. Tungsten (W) can substitute for Mo in nitrate reductase, but the enzyme activity is decreased (Heimer, Wray, and Filner, 1969), as the formation of an active Mo cofactor is prevented. In an experiment with W-treated tobacco (Nicotiana tabacum) plants supplied with N as NOs , Deng, Moureaux, and Caboche (1989) reported a decrease in nitrate reductase activity, but several-fold increases in the accumulation of nitrate reductase apoprotein and corresponding mRNA because of excessive expression of a nitrate reductase structural gene. 

In cyanobacteria, fungi and higher plants with assimilatory nitrate reductase, nitrate enhancement of nitrate reductase levels has been observed. The nitrate-dependent appearance of nitrate reductase is prevented by inhibitors of protein and nucleic acid synthesis (Beevers and Hageman, I%9 Hewitt, 1975). Inhibitor studies indicate that the influence of nitrate is one of induction of protein synthesis rather than an activation of some inert precursor and Zielke and Filner (1971) have demonstrated de novo synthesis of nitrate reductase in response to inducer nitrate. Concurrent degradation of the enzyme was also noted. 

The results can be interpreted according to Cove and Pateman (1969) by a process of autoregulation in which the enzyme is constitutively synthesized at low levels in the absence of nitrate and is induced at high levels in the presence of nitrate. The constitutively formed enzyme is postulated to be partly responsible for its own repression. In the presence of nitrate the enzyme is blocked in its regulatory function and is unable to repress its own synthesis and induction results. Aberrant nitrate reductase is postulated to be unable to repress its own synthesis consequently high aberrant levels are formed in the absence of nitrate. In many respects, this autoregulatory feature is analogous to that reported for the dissimilatory enzyme in bacteria. 

Fig. 1 21]. An ABA concentration as low as 10 M is sufficient to enhance its own metabolism, and this effect can be observed within 2 h of ABA treatment [21]. The formation of the next stable metabolite, DPA, is not enhanced by pretreatments with either ABA or PA [21]. Thus, the enhanced PA formation is unlikely a scavenging mechanism to remove excessive ABA because the tissue would have to enhance the formation of DPA in order to eliminate the biological activities. The self-induction of ABA metabolism can be prevented by transcription and translation inhibitors, suggesting that ABA induces the monooxygenase (or a cofactor for this enzyme) responsible for PA formation. The regulation of ABA metabolism in barley aleurone layers is similar to the induction of nitrate reductase by its substrate, nitrate. In this latter case, treatment of a plant tissue with nitrate enhances its ability to metabolize nitrate. 

Substrate induction is subject to a temporal control of an unknown kind. Substrate induction anddi temporal control of substrate induction of a higher order have been discovered in a few further instances, such as different kinds of nitrate reductase and an enzyme involved in anthocyanin synthesis in Petunia hybrida. In addition, there is quite a number of enzymes for which substrate induction ora temporal control of synthesis of a higher order has been established. We shall become acquainted with examples of the latter type later. 

Various findings suggest that gene material can be activated under the influence of IAA. However, a certain lapse of time, a lag phase, occurs before enzymes are formed after induction by lAA. In the case of the induction of j8-galactosidase of E. coli it amounts to 3-4 minutes, for example. In higher plants the lag phase is longer. In the case of the induction of the nitrate reductase of maize the lag phase was two hours, the shortest found so far in higher plants. 

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