Abstract
Protein–protein interaction networks (interactomes) define the functionality of all biological systems. In apoptosis, proteolysis by caspases is thought to initiate disassembly of protein complexes and cell death. Here we used a quantitative proteomics approach, protein correlation profiling (PCP), to explore changes in cytoplasmic and mitochondrial interactomes in response to apoptosis initiation as a function of caspase activity. We measured the response to initiation of Fas‐mediated apoptosis in 17,991 interactions among 2,779 proteins, comprising the largest dynamic interactome to date. The majority of interactions were unaffected early in apoptosis, but multiple complexes containing known caspase targets were disassembled. Nonetheless, proteome‐wide analysis of proteolytic processing by terminal amine isotopic labeling of substrates (TAILS) revealed little correlation between proteolytic and interactome changes. Our findings show that, in apoptosis, significant interactome alterations occur before and independently of caspase activity. Thus, apoptosis initiation includes a tight program of interactome rearrangement, leading to disassembly of relatively few, select complexes. These early interactome alterations occur independently of cleavage of these protein by caspases.
Highlights
The association of proteins into functional units is a quintessential feature of life, with 150,000–650,000 discrete protein–protein interactions predicted within the human proteome (Hart et al, 2006; Stumpf et al, 2008)
As protein correlation profiling (PCP)-SILAC enables the measurement of cytosolic interactome responses (Kristensen et al, 2012), we reasoned that using a membranecompatible separation method should allow the measurement of organelle/membrane interactome dynamics
We have mapped the cytosolic and organelle membrane interactomes, as well as proteolytic cleavage events, of Jurkat cells undergoing early stages of apoptosis and used this information to address the key assumption in the apoptosis field: that the caspase cascade triggers dissociation of the cellular machinery
Summary
The association of proteins into functional units is a quintessential feature of life, with 150,000–650,000 discrete protein–protein interactions predicted within the human proteome (Hart et al, 2006; Stumpf et al, 2008). The recognition that proteomes are highly interconnected networks, or interactomes, has changed biology’s view of cause and effect, simultaneously highlighting the importance of systems biology and necessitating its use in deciphering complex biological phenomena effects (Barabasi & Oltvai, 2004; Barabasi et al, 2011). This concept of connectivity modulating systems explains how functionally dissimilar systems, such as tissues, can have largely similar compositions (Geiger et al, 2013; Kim et al, 2014; Wilhelm et al, 2014). The very existence of such modules suggests that a network-level view of the proteome is more meaningful for understanding biological states and disease than a detailed picture of specific components (Bandyopadhyay et al, 2010; Barabasi et al, 2011; Califano, 2011)
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