Study of the cell cycle interactome of Arabidopsis thaliana through a targeted proteomics approach

Defining protein complexes is critical to virtually all aspects of cell biology, because many cellular processes are regulated by stable protein complexes and their identification often provides insights into their function. In the introduction of the thesis, an overview is given on proteomic approaches with an emphasis towards study of protein-protein interactions and towards plant proteomic sudies. Next, we describe the development and application of a high-througput tandem affinity purification (TAP)/mass spectrometry platform for cell suspension cultures to analyze protein complexes in Arabidopsis thaliana. The TAP technology was further optimized through the introduction of different alternative TAP tags. Finally, this technology platform was used to map the cell cycle interactome of Arabidopsis thaliana. The resulting protein interaction network among 394 proteins contains 866 interactions, from which 82% has never been described before. Based on a computational analysis, gene networks involved in cell cycle phase transition, DNA replication, and mitosis are presented, together with a list of new candidate cell cycle proteins. This is the first cell cycle interactome for higher eukaryotes mapped by tandem affinity purification of protein comlexes and mass spectromethry analysis. It provides insight in the basic cell cylce machinery and serves as a guide for the investigation of other protein networks by complex purifactinin plants.