Molecular Mechanism Of How Sucrose Transporters Regulate Phloem Loading In Response To Environmental Cues

In a multicellular organism,growth,development,acclimation,and homeostasis require mechanisms that monitor changes in the environment,coordinate reactions in cells and compartments,and adjust transport of ions and metabolites across cellular membranes.All multi-cellular organisms keep a balance between sink and source activities by controlling nutrient transport at strategic positions.Plants depend on a strict regulation of carbon transport to keep the activities of different parts in balance under various environmental conditions.This entails control of the flux of sucrose,the predominant unit of carbon and energy.Sucrose is produced photosynthetically active tissues of the leaf and stem and transported to carbon sink organs in the highly specialized cells of the phloem vascular system.Since transport inside the phloem happens by osmotically-driven mass flow,it is the loading and unloading reactions that determine the transport rate,and depends on the sucrose transporters.In most crops,as well as the model plant Arabidopsis(Arabidopsis thaliana),phloem loading is mediated by secondary active SUCROSE TRANSPORTERs(SUCs or SUTs)after sucrose secretion into the cell wall space by passive SUGARS WILL EVENTUALLY BE EXPORTED TRANSPORTERs(SWEETs)from cells surrounding the phloem.Sucrose transporters(SUTs)play a key role in the allocation and partitioning of photosynthetically fixed carbon in plants,and the transport of sucrose into the phloem vascular tissue by phloem SUTs sets the rate of carbon export from source leaves,like AtSUC2 in Arabidopsis and the AtSUC2-homologues of most crop plants.Despite its strategic role in whole-plant carbon partitioning,little is known about the mechanism of regulation of SUCs responsible for phloem loading.Not knowing the extent to which plants rely on SUT regulation to control carbon partitioning is a major reason for our limited understanding of the control of this process.Here,the question of how SUTs are regulated in response to various stimuli is addressed through a global analysis of publicly available gene expression data in the leaves of five dicots,Arabidopsis,soybean,potato,tomato and poplar,and four monocots,maize,rice,wheat and barley,was investigated firstly.How SUT expression correlates with phloem loading is then further investigated experimentally.Extensive data on expression of SUTs in relation to changes of environmental conditions were obtained through a global analysis of 168 transcriptomics data sets.Results were validated by q PCR measurements and extended by the measurement of photosynthesis rate and phloem sugar content to draw insight on the correlation of SUT expression and sugar export from leaves.For the apoplasmic phloem loaders,a clear difference in transcriptional regulation in response to different environmental stimuli was observed.The consistent patterns of SUT expression under abiotic stress indicates which types of SUTs are involved in the regulation of leaf sugar status and in stress signaling.Furthermore,it is shown that downregulation of phloem loading is likely to be caused by transcriptional regulation of SUTs,while upregulation depends on post-transcriptional regulation.In poplar,expression of Pta SUT4 was found to consistently respond to environmental stimuli,suggesting a significant role in the regulation of sugar export from leaves in this passive symplasmic phloem loader.Gene expression was found to scale with phloem loading in response to certain abiotic stresses,but did not scale with phloem loading in response to increased leaf sugar production.In tomato and Arabidopsis,unchanged expression under high light conditions was compensated by increased protein abundance of the phloem loading SUC,indicating posttranscriptional regulation.Hence,identification and characterization of SUTs-interaction partners using Arabidopsis AtSUC2 as model was further explored.This study is the first that identified interaction partners with regulatory function,investigated their mechanism of regulation and demonstrated its relevance for plant acclimation to different environments.The results revealed that AtSUC2 activity is regulated via its protein turn-over rate and phosphorylation state.The UBIQUITIN-CONJUGATING ENZYME 34(UBC34)was identified as AtSUC2-interaction partner that mediates turn-over in the 26 S proteasome.ubc34 knockout mutants showed increased phloem loading,as well as increased biomass and yield.In contrast,mutants of another AtSUC2-interaction partner,WALL ASSOCIATED KINASE LIKE 8(WAKL8),showed decreased phloem loading and growth.An in vivo assay based on a fluorescent sucrose analogue,confirmed that AtSUC2-phosphorylation by WAKL8 can increase transport activity.Furthermore,this study is the first that investigate the detailed molecular mechanism of AtSUC2 regulation and demonstrate that the manipulation of SUC-regulation can be a viable path to enhance plant biomass.