No abstract

Sugar transporters are central machineries to mediate cross-membrane transport of sugars into the cells, and sugar availability may serve as a signal to regulate the sugar transporters. However, the mechanisms of sugar transport regulation by signal sugar availability remain unclear in plant and animal cells. Here, we report that a sucrose transporter, MdSUT1, and a sorbitol transporter, MdSOT6, both localized to plasma membrane, were identified from apple (Malus domestica) fruit. Using a combination of the split-ubiquitin yeast two-hybrid, immunocoprecipitation, and bimolecular fluorescence complementation assays, the two distinct sugar transporters were shown to interact physically with an apple endoplasmic reticulum-anchored cytochrome b5 MdCYB5 in vitro and in vivo. In the yeast systems, the two different interaction complexes function to upregulate the affinity of the sugar transporters, allowing cells to adapt to sugar starvation. An Arabidopsis (Arabidopsis thaliana) homolog of MdCYB5, AtCYB5-A, also interacts with the two sugar transporters and functions similarly. The point mutations leucine-73 / proline in MdSUT1 and leucine-117 / proline in MdSOT6, disrupting the bimolecular interactions but without significantly affecting the transporter activities, abolish the stimulating effects of the sugar transporter-cytochrome b5 complex on the affinity of the sugar transporters. However, the yeast (Saccharomyces cerevisiae) cytochrome b5 ScCYB5, an additional interacting partner of the two plant sugar transporters, has no function in the regulation of the sugar transporters, indicating that the observed biological functions in the yeast systems are specific to plant cytochrome b5s. These findings suggest a novel mechanism by which the plant cells tailor sugar uptake to the surrounding sugar availability.

Summary• In higher plants, the crosstalk between cold stress responses and reactive oxygen species (ROS) signaling is not well understood.• Two chilling-sensitive mutants, chs4-1 and chs4-3, were characterized genetically and molecularly.• The CHS4 gene, identified by map-based cloning, was found to be identical to LESION SIMULATING DISEASE RESISTANCE 1 (LSD1). We therefore renamed these two alleles lsd1-3 and lsd1-4, respectively. These two mutants exhibited an extensive cell death phenotype under cold stress conditions. Consistently, lsd1-3 plants exposed to cold showed up-regulation of the PR1 and PR2 genes, and increased accumulation of salicylic acid. These results indicate that low temperature is another trigger of cell death in lsd1 mutants. Furthermore, lsd1-3 plants accumulated higher concentrations of H 2 O 2 and total glutathione under cold conditions than wild-type plants. Genetic analysis revealed that PAD4 and EDS1, two key signaling regulators mediating resistance responses, are required for the chilling-sensitive phenotype of lsd1-3.• These findings reveal a role of LSD1 in regulating cell death trigged by cold stress and a link between cold stress responses and ROS-associated signaling.