Modulation of Protein-Interaction States through the Cell Cycle

Abstract

Global profiling of protein expression through the cell cycle has revealed subsets of periodically expressed proteins. However, expression levels alone only give a partial view of the biochemical processes determining cellular events. Using a proteome-wide implementation of the cellular thermal shift assay (CETSA) to study specific cell-cycle phases, we uncover changes of interaction states for more than 750 proteins during the cell cycle. Notably, many protein complexes are modulated in specific cell-cycle phases, reflecting their roles in processes such as DNA replication, chromatin remodeling, transcription, translation, and disintegration of the nuclear envelope. Surprisingly, only small differences in the interaction states were seen between the G1 and the G2 phase, suggesting similar hardwiring of biochemical processes in these two phases. The present work reveals novel molecular details of the cell cycle and establishes proteome-wide CETSA as a new strategy to study modulation of protein-interaction states in intact cells.

Dynamic rewiring of the human interactome by interferon signalling

“…Thermal proximity co-aggregation (TPCA) has demonstrated promise for interrogating protein interaction networks in vivo , or across distinct cellular states [65][66][67] , but to date has been limited to monitoring the dynamics of known interactions or protein complexes, rather than enabling de novo network inference. In contrast, the primary disadvantage associated with SEC-PCP-SILAC is its moderate bias towards proteins of greater cellular abundance [68] .…”

Thermal proteome profiling for interrogating protein interactions

“…More commonly, TPP experiments involve chemical [e.g., drug (Azimi et al, 2018;Becher et al, 2016;Hu et al, 2019;Huber et al, 2015;Kitagawa et al, 2017;Mateus et al, 2018Mateus et al, , 2016Reinhard et al, 2015;Savitski et al, 2014Savitski et al, , 2018 . Some of the perturbations can be applied in a dose-dependent manner (Becher et al, 2016) or time-dependent manner Dai et al, 2018) to improve data analysis or facilitate mechanistic understanding of the perturbation (Fig 2). Using this approach, it has been possible to deconvolute drug targets Huber et al, 2015;Reinhard et al, 2015;Becher et al, 2016;Mateus et al, 2016Mateus et al, , 2018Kitagawa et al, 2017;Azimi et al, 2018;Hu et al, 2019) and enzyme substrates (preprint: Saei et al, 2018), study metabolic shifts Dai et al, 2018;Mateus et al, 2018), or identify protein-protein interactions (Tan et al, 2018).…”

“…Some of the perturbations can be applied in a dose‐dependent manner (Becher et al , ) or time‐dependent manner (Becher et al , ; Dai et al , ) to improve data analysis or facilitate mechanistic understanding of the perturbation (Fig ). Using this approach, it has been possible to deconvolute drug targets (Savitski et al , , ; Huber et al , ; Reinhard et al , ; Becher et al , ; Mateus et al , , ; Kitagawa et al , ; Azimi et al , ; Hu et al , ) and enzyme substrates (preprint: Saei et al , ), study metabolic shifts (Becher et al , ; Dai et al , ; Mateus et al , ), or identify protein–protein interactions (Tan et al , ).…”

“…The next step in a TPP experiment is subjecting the samples to a heat cycle [at a single (Dai et al , ; Franken et al , ) or, more commonly, multiple temperatures], which is generally performed in small volumes in a thermocycler, for rapid and homogenous heat transfer (Fig ). Usually, samples are heated for 3 min, which was initially shown to be sufficient to induce intracellular protein aggregation (Martinez Molina et al , ).…”

“…; or metabolite (preprint:Saei et al, 2018;Dziekan et al, 2019;Sridharan et al, 2019b)], genetic [e.g., gene knock-outBanzhaf et al, 2020)], or enzymatic (preprint:Saei et al, 2018) perturbations; or different cell states [different phase of the cell cycleDai et al, 2018), or growth phase;Fig 2]…”