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.

Elucidation of host-pathogen protein-protein interactions to uncover mechanisms of host cell rewiring.

Infectious diseases are the result of molecular cross-talks between hosts and their pathogens. These cross-talks are in part mediated by host-pathogen protein-protein interactions (HP-PPI). HP-PPI play crucial roles in infections, as they may tilt the balance either in favor of the pathogens' spread or their clearance. The identification of host proteins targeted by viral or bacterial pathogenic proteins necessary for the infection can provide insights into their underlying molecular mechanisms of pathogenicity, and potentially even single out pharmacological intervention targets. Here, we review the available methods to study HP-PPI, with a focus on recent mass spectrometry based methods to decipher bacterial-human infectious diseases and examine their relevance in uncovering host cell rewiring by pathogens.

Systems proteomics approaches to study bacterial pathogens: application to Mycobacterium tuberculosis.

Significant developments and improvements in basic and clinical research notwithstanding, infectious diseases still claim at least 13 million lives annually. Classical research approaches have deciphered many molecular mechanisms underlying infection. Today it is increasingly recognized that multiple molecular mechanisms cooperate to constitute a complex system that is used by a given pathogen to interfere with the biochemical processes of the host. Therefore, systems-level approaches now complement the standard molecular biology techniques to investigate pathogens and their interactions with the human host. Here we review omic studies in Mycobacterium tuberculosis, the causative agent of tuberculosis, with a particular focus on proteomic methods and their application to the bacilli. Likewise, the discussed methods are directly portable to other bacterial pathogens.