EAGER: Defining the phloem-mobile proteins of Arabidopsis

University Of Chicago, Chicago IL

Investigators

Abstract

In higher animals, blood moves throughout the body in veins and arteries, carrying nutrients, hormones and signaling molecules that coordinate the functions of different organs. Plants have an analogous fluid called phloem sap, which moves through phloem vessels to carry sugars from photosynthetic organs to the rest of the plant, and to communicate information about nutritional needs, stress or disease between distant plant organs. Scientists have only a rudimentary understanding of phloem sap-mediated signaling, primarily because it is extremely difficult to collect a phloem sap sample that is not heavily contaminated by fluids from the surrounding cells. While evidence suggests that hundreds of proteins exist in the phloem sap, only a small number are confirmed to actually move and only one has a known function. In this project, the researchers will identify hundreds of phloem sap proteins with confirmed long distance movement. The researcher's novel approach involves grafting two plants with distinct genetic sequences together and then using protein sequence analysis to identify proteins from plant 1 in the phloem vessels of plant 2. If successful, this project will generate the first large dataset of mobile phloem sap proteins, overcoming the current bottleneck in the field of intra-plant signaling. The identity of these proteins will catalyze new research directions and will impact a large number of plant biologists. Phloem signaling is an essential component of the plant's ability to survive biotic and abiotic stresses. Therefore, this project will suggest new strategies for improving crop plant growth and yield. Despite the exciting potential of intra-plant communication via phloem-mobile proteins, there is no current consensus as to the identity of the phloem sap proteome in most plants, including the model plant Arabidopsis thaliana where researchers could best test the function of these proteins. The PI will use a grafting approach to definitively identify proteins that move long distance through a graft union. Taking advantage of the extensive sequence data available for over 1000 Arabidopsis accessions, the PI will identify grafting partners where sequence variation leads to amino acid differences in ~1100 predicted phloem sap proteins. Heterografts will be made, protein samples enriched for phloem sap will be collected from the scion, and Mass Spectrometry will be used to identify rootstock proteins that have travelled to the scion tissue. The proteome generated by this project will rely on observed protein movement. This is in contrast to previous attempts in the field to define the phloem sap proteome based on direct phloem sap extraction, a process that is extremely difficult and plagued with technical difficulties. The definition of hundreds of mobile proteins will overcome the current bottleneck in this field, where the existing phloem sap proteomes are contradictory and generally acknowledged to have large sampling artefacts. Once in hand, the phloem sap proteome will lead to testable hypotheses about phloem sap signaling based on the identity and function of the mobile proteins.