The tonoplast monosaccharide transporter (TMT) family comprises three isoforms in Arabidopsis thaliana, and TMT-green fluorescent protein fusion proteins are targeted to the vacuolar membrane. TMT promoter-b-glucuronidase plants revealed that the TONOPLAST MONOSACCHARIDE TRANSPORTER1 (TMT1) and TMT2 genes exhibit a tissue-and cell type-specific expression pattern, whereas TMT3 is only weakly expressed. TMT1 and TMT2 expression is induced by drought, salt, and cold treatments and by sugar. During cold adaptation, tmt knockout lines accumulated less glucose and fructose compared with wild-type plants, whereas no differences were observed for sucrose. Cold adaptation of wild-type plants substantially promoted glucose uptake into isolated leaf mesophyll vacuoles. Glucose uptake into isolated vacuoles was inhibited by NH 4 þ , fructose, and phlorizin, indicating that transport is energy-dependent and that both glucose and fructose were taken up by the same carrier. Glucose import into vacuoles from two cold-induced tmt1 knockout lines or from triple knockout plants was substantially lower than into corresponding wild-type vacuoles. Monosaccharide feeding into leaf discs revealed the strongest response to sugar in tmt1 knockout lines compared with wild-type plants, suggesting that TMT1 is required for cytosolic glucose homeostasis. Our results indicate that TMT1 is involved in vacuolar monosaccharide transport and plays a major role during stress responses.

The plant cell wall is a dynamic and complex structure whose functional integrity is constantly being monitored and maintained during development and interactions with the environment. In response to cell wall damage (CWD), putatively compensatory responses, such as lignin production, are initiated. In this context, lignin deposition could reinforce the cell wall to maintain functional integrity. Lignin is important for the plant's response to environmental stress, for reinforcement during secondary cell wall formation, and for long-distance water transport. Here, we identify two stages and several components of a genetic network that regulate CWD-induced lignin production in Arabidopsis (Arabidopsis thaliana). During the early stage, calcium and diphenyleneiodonium-sensitive reactive oxygen species (ROS) production are required to induce a secondary ROS burst and jasmonic acid (JA) accumulation. During the second stage, ROS derived from the NADPH oxidase RESPIRATORY BURST OXIDASE HOMOLOG D and JA-isoleucine generated by JASMONIC ACID RESISTANT1, form a negative feedback loop that can repress each other's production. This feedback loop in turn seems to influence lignin accumulation. Our results characterize a genetic network enabling plants to regulate lignin biosynthesis in response to CWD through dynamic interactions between JA and ROS.

SUMMARYThe vacuolar membrane is involved in solute uptake into and release from the vacuole, which is the largest plant organelle. In addition to inorganic ions and metabolites, large quantities of protons and sugars are shuttled across this membrane. Current models suggest that the proton gradient across the membrane drives the accumulation and/or release of sugars. Recent studies have associated AtSUC4 with the vacuolar membrane. Some members of the SUC family are plasma membrane proton/sucrose symporters. In addition, the sugar transporters TMT1 and TMT2, which are localized to the vacuolar membrane, have been suggested to function in proton-driven glucose antiport. Here we used the patch-clamp technique to monitor carriermediated sucrose transport by AtSUC4 and AtTMTs in intact Arabidopsis thaliana mesophyll vacuoles. In the whole-vacuole configuration with wild-type material, cytosolic sucrose-induced proton currents were associated with a proton/sucrose antiport mechanism. To identify the related transporter on one hand, and to enable the recording of symporter-mediated currents on the other hand, we electrophysiologically characterized vacuolar proteins recognized by Arabidopsis mutants of partially impaired sugar compartmentation. To our surprise, the intrinsic sucrose/proton antiporter activity was greatly reduced when vacuoles were isolated from plants lacking the monosaccharide transporter AtTMT1/TMT2. Transient expression of AtSUC4 in this mutant background resulted in proton/sucrose symport activity. From these studies, we conclude that, in the natural environment within the Arabidopsis cell, AtSUC4 most likely catalyses protoncoupled sucrose export from the vacuole. However, TMT1/2 probably represents a proton-coupled antiporter capable of high-capacity loading of glucose and sucrose into the vacuole.