Tierney, Mary Role of endocytic pathways in controlling cell wall structure
National Institute of Food and Agriculture
The purpose of this project is to characterize genes important in regulating the dynamic structure of the plant cell wall during growth. Previously, we characterized an endosomal trafficking pathway defined by the SNARE VTI13, whose transcript level is regulated by cell wall structure and whose function is essential for the secretion or assembly of cell wall components and polarized growth in Arabidopsis. During the past year we have extended these studies by characterizing VPS26C, a gene that shares a pathway with VTI13 and is also required for polarized growth and cell wall organization.
We have shown that vps26c mutant alleles are defective in root hair growth mannitol or NaCl is added to the media. We have also shown that cell wall organization in the vps26c mutants is altered in roots and root hairs. These phenotypes are complemented when we introduce a gene encoding a GFP-VPS26C fusion protein into the mutant background, confirming that the phenotypes we observed in the vsp26c mutants are due to a loss of VPS26C function. Lastly, we have generated a vti13vps26c double mutant and shown that these plants have normal root hair growth and wall organization, indicating vps26c is a genetic suppressor of vti13 in arabidopsis. These studies define two proteins that function in a common cellular trafficking pathway required for growth and cell wall organization in plant roots.
To investigate whether VPS26C orthologs in other plant species have a conserved function, we constructed a phylogeny using available genome and transcript databases. These studies indicated that orthologs of VPS26C are present in many eudicot species. In addition, work from other labs has shown that VPS26C is part of a small clade containing both plant and animal sequences. We are currently examining whether non-plant VPS26C orthologs (human) as well as plant VPS26C orthologs (poplar) will complement the vps26c mutant root hair phenotype. This approach represents a first step in defining whether the cellular trafficking pathway involving VPS26C is conserved in both animals and plants. We are also investigating the proteins that VPS26C interacts with in plants using both genetic approaches and Biomolecular Fluorescence Complementation studies.
This work has been disseminated to the plant biology community by oral and poster presentations at two meetings this year – the 5th PanAmerican Plant Membrane Workshop and the Plant Cell Wall Gordon Conference. In the upcoming year, we will investigate three additional questions. The first is to identify the protein cargo that VPS26C recycles to the Golgi or trans-Golgi. We have established a collaboration with Dr. Federica Brandizzi at the DOE labs at Michigan State University where we will screen a yeast two hybrid library for VPS26C interacting proteins. Secondly, in collaboration with Dr. Michael Hahn’s lab at the CCRC in Georgia, we will use glycome profiling to characterize changes in cell wall structure in the vps26c and vti13 that may be responsible for altered root hair growth in these mutants. Lastly we will begin a genetic suppressor screen for vps26c mutants. These studies will define additional components of a conserved, cellular pathway in plants required for wall organization critical for growth under conditions of salt and/or drought stress.