Rapid Profiling of Protein Complex Reorganization in Perturbed Systems

Protein complexes constitute the primary functional modules
of
cellular activity. To respond to perturbations, complexes undergo
changes in their abundance, subunit composition, or state of modification.
Understanding the function of biological systems requires global strategies
to capture this contextual state information. Methods based on cofractionation
paired with mass spectrometry have demonstrated the capability for
deep biological insight, but the scope of studies using this approach
has been limited by the large measurement time per biological sample
and challenges with data analysis. There has been little uptake of
this strategy into the broader life science community despite its
rich biological information content. We present a rapid integrated
experimental and computational workflow to assess the reorganization
of protein complexes across multiple cellular states. The workflow
combines short gradient chromatography and DIA/SWATH mass spectrometry
with a data analysis toolset to quantify changes in a complex organization.
We applied the workflow to study the global protein complex rearrangements
of THP-1 cells undergoing monocyte to macrophage differentiation and
subsequent stimulation of macrophage cells with lipopolysaccharide.
We observed substantial proteome reorganization on differentiation
and less pronounced changes in macrophage stimulation. We establish
our integrated differential pipeline for rapid and state-specific
profiling of protein complex organization.