Pervasive Protein Thermal Stability Variation during the Cell Cycle

摘要

class="head no_bottom_margin" id="sec1title">IntroductionMass spectrometry (MS)-based proteomics is a powerful method to accurately quantify distinct features of proteomes () and has provided comprehensive insight into the variability of protein abundance under distinct biological conditions, including different cell types (), protein complexes (), and subcellular fractions (). Recent advances in MS multiplexing technologies () enable the global measurement of protein melting curves (thermal proteome profiling; TPP) (). TPP and a partial digestion-based methodology for assessing protein thermal stability () add another dimension to MS-based proteomics to quantify various states of the proteome. TPP can detect individual proteins that are stabilized in situ by binding to a ligand and, therefore, is useful for profiling drug targets and off-targets (, ). However, its utility to uncover global changes in protein thermal stability in distinct biological contexts is unclear.The eukaryotic cell cycle is the key regulatory circuit that controls the temporal separation of fundamental processes that facilitate cell proliferation. It is well established that various aspects of proteome organization, including protein abundance and post-translational modifications, vary during cell-cycle progression (, ). We hypothesized that cell-cycle-dependent post-translational modifications, protein-protein interactions, and spatial rearrangements to distinct biophysical environments globally influence protein thermal stability (, , , ). Here, we systematically measured in situ protein thermal stability, abundance, and solubility during cell-cycle progression on a proteome-wide scale.We report the pervasive variation of protein thermal stability during the cell cycle and link it to various biological processes including transcription, spindle formation and key metabolic pathways. Further, intrinsically disordered proteins are stabilized during mitosis, coinciding with fundamental rearrangements of the proteome and the spatial outline of the cell. These changes coincide with extensive sumoylation and mitotic phosphorylation, suggesting that post-translational modifications might promote thermal stability and, in turn, prevent protein aggregation during mitotic spindle formation and chromosomal separation. Protein stabilization serves as a proxy for biological activity and complex formation, thereby revealing new players in the cell cycle. Our comprehensive analysis of cell-cycle-dependent variation of protein thermal stability, abundance, and solubility provides a valuable resource to advance the fields of transcription, structural biology, intrinsically disordered proteins, metabolism, and the cell cycle.