A CO2-Flux Mechanism Operating via pH-Polarity in Hydrilla Verticillata Leaves With C3 and C4 Photosynthesis

The aquatic angiosperm Hydrilla verticillata lacks Kranz anatomy, but has an inducible, C₄-based, CO₂ concentrating mechanism (CCM) that concentrates CO₂ in the chloroplasts. Both C₃ and C₄ Hydrilla leaves showed light-dependent pH polarity that was suppressed by high dissolved inorganic carbon (DIC). At low DIC (0.25 mol m⁻³), pH values in the unstirred water layer on the abaxial and adaxial sides of the leaf were 4.2 and10.3, respectively. Abaxial apoplastic acidification served as a CO₂ flux mechanism (CFM), making HCO₃ ⁻ available for photosynthesis by conversion to CO₂. DIC at 10 mol m⁻³ completely suppressed acidification and alkalization. The data, along with previous results, indicated that inhibition was specific to DIC, and not a buffer effect. Acidification and alkalization did not necessarily show 1:1 stoichiometry; their kinetics for the apolar induction phase differed, and alkalization was less inhibited by 2.5 mol m⁻³ DIC. At low irradiance (50 μmol photons m⁻² s⁻¹), where CCM activity in C₄ leaves is minimized, both leaf types had similar DIC inhibition of pH polarity. However, as irradiance increased, DIC inhibition of C₃ leaves decreased. In C₄ leaves the CFM and CCM seemed to compete for photosynthetic ATP and/or reducing power. The CFM may require less, as at low irradiance it still operated maximally, if DIC] was low. Iodoacetamide (IA), which inhibits CO₂ fixation in Hydrilla, also suppressed acidification and alkalization, especially in C₄ leaves. IA does not inhibit the C₄ CCM, which suggests that the CFM and CCM can operate independently. It has been hypothesized that irradiance and DIC regulate pH polarity by altering the chloroplastic DIC], which effects the chloroplast redox state and subsequently redox regulation of a plasma-membrane H⁺-ATPase. The results lend partial support to a down-regulatory role for high chloroplastic DIC], but do not exclude other sites of DIC action. IA inhibition of pH polarity seems inconsistent with the chloroplast NADPH/NADP⁺ ratio being the redox transducer. The possibility that malate and oxaloacetate shuttling plays a role in CFM regulation requires further investigation. This revised version was published online in June 2006 with corrections to the Cover Date.