Manipulation of ethanol production in anoxic rice coleoptiles by exogenous glucose determines rates of ion fluxes and provides estimates of energy requirements for cell maintenance during anoxia

Abstract

Ethanol production by anoxic, excised, 7-10 mm tips of rice coleoptiles was manipulated using a range of exogenous glucose concentrations. Such a dose-response curve enabled good estimates at which level of ethanol production (and hence by inference ATP production), injury commenced and also allowed assessments of energy requirements for maintenance in anoxia. Rates of net uptake or loss of K+ and P by these excised coleoptile tips were related to rates of ethanol production (r2 of 0.59 and 0.68, respectively). At 72 h anoxia, ATP levels in excised tips were similar at 0, 2.5, and 50 mol m(-3) exogenous glucose, despite large differences in the inferred rates of ATP production. At 96 h anoxia, tips without exogenous glucose had low ATP concentrations; these may be the cause or the consequence of cell injury. In tips without glucose, injury was indicated by losses of K+ and Cl- between 72-96 h anoxia, and during the first hour after re-aeration, while later than 1 h after re-aeration, rates of net uptake were substantially lower than for re-aerated tips previously in anoxia with exogenous glucose. Between 96 h and 124 h anoxia, ion losses from tips without exogenous glucose increased while recovery of net uptake after re-aeration was very sluggish and incomplete. The energy requirement for maintenance of health and survival of anoxic coleoptile tips, expressed on a fresh weight basis, was lower than for three other anoxia-tolerant plant tissues/cells, studied previously. However, the energy requirement on a protein basis was assessed at 1.4 micromol ATP mg(-1) protein h(-1) and this value is 2.6-5.4-fold higher than for the other plant tissues/cells. Yet, this requirement was still only 58-88% of the published values for aerated tissues. The reason for this relatively high ATP requirement per unit protein in anoxic rice coleoptiles remains to be elucidated.

Energetics of acclimation to NaCl by submerged, anoxic rice seedlings

“…4A) supports the suggestions by Colmer et al (2001) and Huang et al (2005) that under anoxia without additional adverse factors, rice coleoptiles down-regulate ethanol formation, once acclimation with its high energy demands is completed. The acclimative advantage of such down-regulation of ethanolic fermentation would be to prolong the availability of substrates needed to survive extended periods of anoxia (cf.…”

“…Experiments were either with intact seedlings (seed and coleoptile) or with excised coleoptile tips (8-12 mm in length, see Huang et al, 2005), dependent on the purpose of the experiment. Both the seedlings for use intact and the seedlings grown to excise coleoptiles were always submerged first in nutrient solution continuously bubbled with air for 36 h, then an N 2 /air mix so that dissolved O 2 was 0Á06 mM (hypoxia) for 30 h, followed by imposition of anoxia, in the dark at 30 C. The period of hypoxia (i.e.…”

“…This 'healing' is required because during the first 1-2 h after excision there were K þ losses from rice coleoptiles (Colmer et al, 2001). Glucose in the nutrient solution was then changed to 5 mM for the aerated and 50 mM for the anoxic treatments with excised coleoptiles (see below); these concentrations prevented excessive sugar accumulation in aerated tissues and ensured adequate glucose uptake in anoxic tissues (Huang et al, 2005).…”

“…† In anoxia it is assumed that the energy requirement for acclimation to NaCl has to be added to the requirement for 'basic' cell maintenance of 3Á3 lmol g À1 f. wt h À1 established under benign conditions (Huang et al, 2005). ATP is also diverted from growth, at 50 mM NaCl growth is reduced by 75 % (Fig.…”

“…The alternative of producing organic solutes, rather than using ions as osmolytes, is about one order of magnitude more energy-expensive than the use of ions (bacteria, Over, 1999; vascular plants, Raven, 1985). Such energy costs may become too taxing in anoxic tissues, which judged on ethanol formation by anoxic rice coleoptiles have rates of glycolysis yielding 7-10 % of the ATP formed by tissues in air with oxidative phosphorylation (Gibbs and Greenway, 2003;Huang et al, 2005).…”