Intrinsic characteristics of the proton pump in the luminal membrane of a tight urinary epithelium. The relation between transport rate and delta mu H.

A number of tight urinary epithelia, as exemplified by the turtle bladder, acidify the luminal solution by active transport of H+ across the luminal cell membrane. The rate of active H+ transport (JH) decreases as the electrochemical potential difference for H+ [delta mu H = mu H(lumen) - mu H(serosa)] across the epithelium is increased. The luminal cell membrane has a low permeability for H+ equivalents and a high electrical resistance compared with the basolateral cell membrane. Changes in JH thus reflect changes in active H+ transport across the luminal membrane. To examine the control of JH by delta mu H in the turtle bladder, transepithelial electrical potential differences (delta psi) were imposed at constant acid-base conditions or the luminal pH was varied at delta psi = 0 and constant serosal PCO2 and pH. When the luminal compartment was acidified from pH 7 to 4 or was made electrically positive, JH decreased as a linear function of delta mu H as previously described. When the luminal compartment was made alkaline from pH 7 to 9 or was made electrically negative, JH reached a maximal value, which was the same whether the delta mu H was imposed as a delta pH or a delta psi. The nonlinear JH vs. delta mu H relation does not result from changes in the number of pumps in the luminal membrane or from changes in the intracellular pH, but is a characteristic of the H+ pumps themselves. We propose a general scheme, which, because of its structural features, can account for the nonlinearity of the JH vs. delta mu H relations and, more specifically, for the kinetic equivalence of the effects of the chemical and electrical components of delta mu H. According to this model, the pump complex consists of two components: a catalytic unit at the cytoplasmic side of the luminal membrane, which mediates the ATP-driven H+ translocation, and a transmembrane channel, which mediates the transfer of H+ from the catalytic unit to the luminal solution. These two components may be linked through a buffer compartment for H+ (an antechamber).

Enhanced Cold Tolerance in Transgenic Tobacco Expressing a Chloroplast Omega-3 Fatty Acid Desaturase Gene under the Control of a Cold-inducible Promoter

A new cold-inducible genetic construct was cloned using a chloroplast-specific omega-3-fatty acid desaturase gene (FAD7) under the control of a cold-inducible promoter (cor15a) from Arabidopsis thaliana. RT-PCR confirmed a marked increase in FAD7 expression, in young Nicotiana tabacum (cv. Havana) plants harboring cor15a-FAD7, after a short-term exposure to cold. When young, cold-induced tobacco seedlings were exposed to low-temperature (0.5, 2 or 3.5 degrees C) for up to 44 days, survival within independent cor15a-FAD7 transgenic lines (40.2-96%) was far superior to the wild type (6.7-10.2%). In addition, the major trienoic fatty acid species remained stable in cold-induced cor15a-FAD7 N. tabacum plants under prolonged cold storage while the levels of hexadecatrienoic acid (16:3) and octadecatrienoic acid (18:3) declined in wild type plants under the same conditions (79 and 20.7% respectively). Electron microscopy showed that chloroplast membrane ultrastructure in cor15a-FAD7 transgenic plants was unaffected by prolonged exposure to cold temperatures. In contrast, wild type plants experienced a loss of granal stacking and disorganization of the thylakoid membrane under the same conditions. Changes in membrane integrity coincided with a precipitous decline in leaf chlorophyll concentration and low survival rates in wild type plants. Cold-induced double transgenic N. alata (cv. Domino Mix) plants, harboring both the cor15a-FAD7 cold-tolerance gene and a cor15a-IPT dark-tolerance gene, exhibited dramatically higher survival rates (89-90%) than wild type plants (2%) under prolonged cold storage under dark conditions (2 degrees C for 50 days).