pH regulation in anoxic rice coleoptiles at pH 3.5: biochemical pHstats and net H+ influx in the absence and presence of NO3−

Abstract

During anoxia, cytoplasmic pH regulation is crucial. Mechanisms of pH regulation were studied in the coleoptile of rice exposed to anoxia and pH 3.5, resulting in H+ influx. Germinating rice seedlings survived a combination of anoxia and exposure to pH 3.5 for at least 4 d, although development was retarded and net K+ efflux was continuous. Further experiments used excised coleoptile tips (7–10 mm) in anoxia at pH 6.5 or 3.5, either without or with 0.2 mM NO3−, which distinguished two processes involved in pH regulation. Net H+ influx (μmol g−1 fresh weight h−1) for coleoptiles with NO3− was ∼1.55 over the first 24 h, being about twice that in the absence of NO3−, but then decreased to 0.5–0.9 as net NO3− uptake declined from ∼1.3 to 0.5, indicating reduced uptake via H+–NO3− symports. NO3− reduction presumably functioned as a biochemical pHstat. A second biochemical pHstat consisted of malate and succinate, and their concentrations decreased substantially with time after exposure to pH 3.5. In anoxic coleoptiles, K+ balancing the organic anions was effluxed to the medium as organic anions declined, and this efflux rate was independent of NO3− supply. Thus, biochemical pHstats and reduced net H+ influx across the plasma membrane are important features contributing to pH regulation in anoxia-tolerant rice coleoptiles at pH 3.5.

Energetics of acclimation to NaCl by submerged, anoxic rice seedlings

“…5C). Furthermore, excised coleoptiles at 0Á3 mM NaCl also accumulated Cl À at 0Á5 lmol g À1 f. wt h À1 between 60 and 84 h of anoxia (Greenway et al, 2012), although this did not occur in the present experiments (Fig. 6B).…”

“…Importantly, in the coleoptile of anoxic rice seedlings, Na þ and Cl À as high as 140 mM can function as 'cheap' osmolytes. Consistently, anoxic rice coleoptiles can transport Cl À against its free energy gradient (Zhang and Greenway, 1995;Greenway et al, 2012).…”

“…For intact seedlings, the most plausible cause for the growth reduction when anoxic and combined with 100 mM NaCl, compared with 0Á3 mM NaCl, is that under anoxia processes essential for survival have priority for energy resources over growth (Greenway et al, 2012;Atwell et al, 2015). The energetic costs of sustaining ion fluxes and cellular compartmentation are presumably the main cause of the poorer growth of anoxic seedlings at 100 mM than at 0Á3 and 50 mM NaCl.…”

“…For interpretation of ion fluxes and energy formation by the coleoptile, excision of this organ and provision of exogenous glucose provides an anoxia-tolerant system for measurements of ion fluxes and ethanol formation (e.g. these excised tissues survive more than 100 h of anoxia, Greenway et al, 2012).…”

Tolerance of anaerobic conditions caused by flooding during germination and early growth in rice (Oryza sativa L.)

“…Reggiani et al (1993, 1995) suggested that nitrate is assimilated in rice seeds germinating under anaerobic conditions. This hypothesis was supported by the studies of Fan et al (1997) in rice coleoptiles and recently, Greenway et al (2012) reported that nitrate is converted to ammonium in anoxic rice coleoptiles, which would maintain the pHstat. However, nitrate is quickly reduced in anaerobic soils and its implication in the internal pH balance of rice under anoxic stress is not clear (Greenway et al, 2012).…”