Hypoxic pretreatment (3 kPa oxygen) of maize (Zea mays L.) root tips improved their survival time in a subsequent anoxic incubation from 10 h to more than 3 d, provided that glucose was added to the medium to sustain metabolism. The glycolytic flux (lactate + ethanol) was the same in both pretreated and untreated root tips during the 1st h after transfer to anoxia. It was only after 2 h that it declined sharply in untreated tips, but was sustained in pretreated ones. Right after the transition from normoxia to anoxia of untreated root tips, the only fermentative product detected was lactic acid, which accumulated in a 7:1 proportion after 30 min in tissue and medium, respectively. It took 10 min before ethanol could be detected and 20 min for it to be produced at its maximum rate at the expense of lactate production, which slowed down. In contrast, in hypoxically pretreated root tips, ethanol was produced at a maximum rate right after the transfer to anoxia. Concurrently, low amounts of lactic acid were produced that accumulated in a 1:1 proportion after 30 min in tissue and medium, respectively. This large efflux of lactic acid could account for the higher cytoplasmic pH values always found in pretreated tissues. The presence of cycloheximide during pretreatment abolished this difference, suggesting that the greater efficiency of lactate efflux was linked to protein synthesis. The role of lactate in cytosolic pH regulation and in sensitivity to anoxia is discussed.Most mesophytes, including maize, are intolerant to anoxic conditions. However, pretreatment at low oxygen pressure induces mechanisms that improve their survival capacity under subsequent anoxic conditions. These mechanisms lead to two kinds of survival strategies. One consists of the formation of lysigenous cortical aerenchymas, helping to maintain a high respiration rate in the tissues by improving internal oxygen transport (5). This is a slow (days) and nonreversible mechanism induced by ethylene (4). The second strategy consists of the metabolic acclimation that apparently improves the energy relationships of anoxic tissues and contributes to a more prolonged survival in the absence of oxygen (9,25). This response is fast (hours) and involves the induction of a limited number of 'anaerobic polypeptides' (23). These polypeptides are different from the 'heat shock proteins ' (14), and those that have been identified are enzymes involved in the glycolytic and fermentative pathways (18,19,24). However, the significance of these proteins in tolerance to anoxia has rarely been examined and remains unclear. The results obtained by Hanson and colleagues (6, 7) on hypoxically inducible LDH' in barley roots and aleurone tissues imply some unrecognized function for this enzyme. Furthermore, there is evidence that ADH levels, which are known to increase dramatically during hypoxic acclimation, are not correlated with the better survival capacity of acclimated maize root tips in anoxia (22).Metabolic studies of survival of plant tissues in the absence...

We tested the hypothesis (J.-H. Xia and P.H. Saglio 119921 Plant Physiol 1 0 0 40-46) that the enhanced ability of maize (Zea mays) root tips to survive anoxia, elicited by a 4-h exposure to 3% O2 ("acclimation"), is due to less cytoplasmic acidosis early in anoxia. Cytoplasmic pH and fermentation readions were monitored in excised and intad (attached) maize root tips by simultaneous in vivo 13C-and "P-NMR spectroscopy. We demonstrate that both excised and intad acclimated root tips have significantly higher cytoplasmic pH values under anoxia. This reduction in cytoplasmic acidosis is greater in intact root tips. Remarkably, cytoplasmic pH does not change when root tips are transferred from 3% O , to anoxia. The earlier observation of considerable ladate efflux and lowered intracellular lactate in excised, acclimated root tips (ibid.) was extended to intact seedlings. The predominant fermentation end product retained in the cells of acclimated root tips is alanine. We discuss the relationship between cytoplasmic pH and levels of intracellular lactate and alanine in sugar-replete roots, and the role of cytoplasmic pH in determining survival under anoxia.

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To investigate the biochemical response of freshwater green algae to elevated CO 2 concentrations, Chlorella pyrenoidosa Chick and Chlamydomonas reinhardtii Dang cells were cultured at different CO 2 concentrations within the range 3-186 µmol/L and the biochemical composition, carbonic anhydrase (CA), and nitrate reductase activities of the cells were investigated. Chlorophylls (Chl), carotenoids, carbonhydrate, and protein contents were enhanced to varying extents with increasing CO 2 concentration from 3-186 µmol/L. The CO 2 enrichment significantly increased the Chl a/Chl b ratio in Chlorella pyrenoidosa, but not in Chlamydomonas reinhardtii. The CO 2 concentration had significant effects on CA and nitrate reductase activity. Elevating CO 2 concentration to 186 µmol/L caused a decline in intracellular and extracellullar CA activity. Nitrate reductase activity, under either light or dark conditions, in C. reinhardtii and C. pyrenoidosa was also significantly decreased with CO 2 enrichment. From this study, it can be concluded that CO 2 enrichment can affect biochemical composition, CA, and nitrate reductase activity, and that the biochemical response was species dependent.

We tested the hypothesis that ATP levels and energy charge determine the resistance of maize (Zea mays) root tips to anoxia. We focused on root tips of whole maize seedlings that had been acclimated to low O , by exposure to an atmosphere of 3% (v/v) O, in N,. Acclimated anoxic root tips characteristically have higher ATP levels and energy charge and survive longer under anoxia than nonacclimated tips. We poisoned intact, acclimated root tips with either fluoride or mannose, causing decreases in ATP and energy charge to values similar to or, i n most cases, below those found in nonacclimated anoxic tips. with the exception of the highest fluoride concentration used, the poisoned, acclimated tips remained much more tolerant of anoxia than nonacclimated root tips. We conclude that high ATP and energy charge are not components critical for the survival of acclimated root tips during anoxia. The reduced nucleotide status in poisoned, acclimated root tips had little effect on cytoplasmic pH regulation during anoxia. This result indicates that i n anoxic, acclimated root tips either cytoplasmic pH regulation is not dominated by ATP-dependent processes or these processes can continue i n vivo largely independently of any changes i n ATP levels i n the physiological range. The role of glycolytic flux in survival under anoxia is discussed.Evaluation of the relative importance of biochemical parameters as determinants of plant performance under environmental stresses, such as low O,, is complicated by the fact that stresses disturb whole networks of biochemical processes (reviewed by Kennedy et al., 1992;Perata and Alpi, 1993;Ricard et al., 1994). Many observed differences between stressed and unstressed, or tolerant and intolerant, plants may well be manifestations of this general disturbance rather than revelations of fundamental processes responsible for the behavior of a particular plant in different environments.The primary biochemical effect of low O, on plant cells is an inhibition of respiratory ATP synthesis, leading to a switch to a lower overall rate of ATP synthesis via fermentation (Beevers, 1961). Given the central role of ATP in cell function (Stryer, 19881, the "energy crisis" in plant cells evoked by low O, would appear to be the direct cause of the eventual loss of function. The resistance of maize root tips to anoxia can be enhanced by pretreatment of intact seedlings with hypoxia ( 2 4 % O,) (Saglio et al., 1988;Johnson et al., 1989). Associated with this acclimation response is the ability to maintain a high glycolytic rate during long periods of anoxia (Saglio et al., 1988;Hole et al., 1992;Xia and Saglio, 1992) and higher ATP levels and energy charge (Saglio et al., 1988;Johnson et al., 1994). One straightforward interpretation of this phenomenon is that the primary acclimation response leading to improved tolerance of anoxia is the enhanced energy metabolism, which relieves the shortage of ATP and facilitates cell function. Moreover, reported differences between acclimated and nonacclimated root t...