Research Training Group 2498

Research Training Group 2498

Communication and Dynamics of Plant Cell Compartments

Projects


LC-MS based mapping of organellar protein trafficking and turn-over

Project Leader// Dr. Wolfgang Hoehenwarter

Graduate// Mohamed Ayash

Leibniz Institute of Plant Biochemistry
Proteome Analytics


Proteins perform different tasks at different sub-cellular locations, and communication and molecular trafficking between the organelles is essential for coordinated activity and cell vitality. The subcellular compartments wherein the proteins perform their various functions can be distinct from the sites of protein synthesis. Therefore, direct measurements of protein abundance – as opposed to measurements of cognate transcripts – are explicit. In recent years progress in fractionation and liquid chromatography/mass spectrometry (LC-MS), live-cell tagging and LC-MS and immuno-fluorescence approaches have greatly advanced spatial proteomics for sub-cellular mapping of protein abundance and function. Here we aim to apply these new methods to the proteomic analysis of plant organelles. In the past, we have conducted considerable sub- cellular fractionation, nuclei isolation and nuclear proteomics experiments in Arabidopsis. We now want to direct our activities primarily towards mapping the spatial and temporal dynamics of the nuclear proteome. In addition we will expand these efforts towards the plastid proteome in light of the diverse molecular interactions between these compartments studied in many of the other projects in the RTG. We aim to understand communication, in particular protein trafficking, synthesis and degradation in protein shuttling between these organelles in the context of plant stress. We thus provide expert discovery and targeted proteomics applications on our Orbitrap Velos Pro and QExactive Plus mass spectrometers in collaborative projects of the RTG. Research in this project is subdivided in the following work packages:

  1. Isolation of nuclei and plastids
  2. In vivo labeling of nuclear proteins and LC-MS
  3. In vivo protein pulse labeling and protein turn-over rate measurement by LC-MS
  4. Application to plant stress responses