When chilling-sensitive plants are chilled, root hydraulic conductance (L o ) declines precipitously; L o also declines in chillingtolerant plants, but it subsequently recovers, whereas in chilling-sensitive plants it does not. As a result, the chilling-sensitive plants dry out and may die. Using a chilling-sensitive and a chilling-tolerant maize genotype we investigated the effect of chilling on L o , and its relationship to osmotic water permeability of isolated root cortex protoplasts, aquaporin gene expression, aquaporin abundance, and aquaporin phosphorylation, hydrogen peroxide (H 2 O 2 ) accumulation in the roots and electrolyte leakage from the roots. Because chilling can cause H 2 O 2 accumulation we also determined the effects of a short H 2 O 2 treatment of the roots and examined the same parameters. We conclude from these studies that the recovery of L o during chilling in the chilling-tolerant genotype is made possible by avoiding or repairing membrane damage and by a greater abundance and/or activity of aquaporins. The same changes in aquaporins take place in the chilling-sensitive genotype, but we postulate that membrane damage prevents the L o recovery. It appears that the aquaporin response is necessary but not sufficient to respond to chilling injury. The plant must also be able to avoid the oxidative damage that accompanies chilling.Many crop species such as maize (Zea mays) that originate in tropical regions are sensitive to low temperatures and grow poorly after exposure to a cold period (Miedema, 1982). Below 12°C to 14°C, maize seedlings are damaged as a result of leaf dehydration and an inhibition of photosynthesis. Such damage may result in the plant's death (Miedema, 1982). With the wide utilization in temperate climates of crops of tropical origin, the adverse effect of chilling is an important agricultural problem the physiological basis of which can be elucidated. Cold-tolerant species increase their root hydraulic conductance (L o ) during low temperature periods (Fennell and Markhart, 1998;Bigot and Boucaud, 2000), and the same phenomenon has been observed in a chilling-tolerant genotype of maize (Aroca et al., 2001b(Aroca et al., , 2003b. The water deficit in the aerial parts of a chilling-sensitive maize genotype is caused in part by low temperature inhibition of L o (Aroca et al., 2001b(Aroca et al., , 2003b. The recovery of L o in cold-tolerant plants may be associated with an increase in the activity or the abundance of aquaporins in the plasma membranes of root cells. On the other hand, the opposite may happen in cold-sensitive species: aquaporins may be down-regulated or inactivated (Aroca et al., 2001b;Sanders and Markhart, 2001;). Lee et al. (2004b) recently showed that in cucumber (Cucumis sativus), a cold sensitive species, a brief exposure to low temperature reduces root pressure, hydraulic conductivity, and active nutrient transport. These authors also postulated that changes in the activity of aquaporins underlie the changes in hydraulic conductivity.Response to cold,...

MEC-4 and MEC-10 are the pore-forming subunits of the sensory mechanotransduction complex that mediates touch sensation in Caenorhabditis elegans (O'Hagan, R., M. Chalfie, and M.B. Goodman. 2005. Nat. Neurosci. 8:43–50). They are members of a large family of ion channel proteins, collectively termed DEG/ENaCs, which are expressed in epithelial cells and neurons. In Xenopus oocytes, MEC-4 can assemble into homomeric channels and coassemble with MEC-10 into heteromeric channels (Goodman, M.B., G.G. Ernstrom, D.S. Chelur, R. O'Hagan, C.A. Yao, and M. Chalfie. 2002. Nature. 415:1039–1042). To gain insight into the structure–function principles that govern gating and drug block, we analyzed the effect of gain-of-function mutations using a combination of two-electrode voltage clamp, single-channel recording, and outside-out macropatches. We found that mutation of A713, the d or degeneration position, to residues larger than cysteine increased macroscopic current, open probability, and open times in homomeric channels, suggesting that bulky residues at this position stabilize open states. Wild-type MEC-10 partially suppressed the effect of such mutations on macroscopic current, suggesting that subunit–subunit interactions regulate open probability. Additional support for this idea is derived from an analysis of macroscopic currents carried by single-mutant and double-mutant heteromeric channels. We also examined blockade by the diuretic amiloride and two related compounds. We found that mutation of A713 to threonine, glycine, or aspartate decreased the affinity of homomeric channels for amiloride. Unlike the increase in open probability, this effect was not related to size of the amino acid side chain, indicating that mutation at this site alters antagonist binding by an independent mechanism. Finally, we present evidence that amiloride block is diffusion limited in DEG/ENaC channels, suggesting that variations in amiloride affinity result from variations in binding energy as opposed to accessibility. We conclude that the d position is part of a key region in the channel functionally and structurally, possibly representing the beginning of a pore-forming domain.

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