Identification of cell cycle dependent interaction partners of the septins by quantitative mass spectrometry

The septins are a conserved family of GTP-binding proteins that, in the baker's yeast, assemble into a highly ordered array of filaments at the mother bud neck. These filaments undergo dramatic structural rearrangements during the cell cycle which are mechanistically not yet understood. We aimed at identifying key components that are involved in the transitions of the septins. Combining cell synchronization and quantitative affinity-purification mass-spectrometry (AP-MS), we identified interaction partners of the septins at three distinct stages of the cell cycle. A total of 83 interaction partners of the septins were assigned. We detected DNA-interacting/nuclear proteins and proteins involved in ribosome biogenesis and protein synthesis predominantly present in G1 arrested cells without an assembled septin structure. In addition, two distinct sets of regulatory proteins that are specific for cells at S-phase with a stable septin collar or at mitosis with split septin rings were identified. Complementary methods such as the SPLIFF method allowed us to more exactly define the spatial and temporal characteristics of selected candidate proteins of the AP-MS screen.