MAPPIT toolbox

Protein interactions are essential in virtually all processes within the cell. Current estimates suggest that the human genome contains approximately 20.000 protein-encoding genes, leading to a staggering number of possible interactions. Moreover, it is well known that the complexity of the proteome extends well beyond the genome due to phenomena such as alternative splicing and secondary modifications, further expanding both the size and complexity of the human interactome.

Because much of the biology of a cell is governed by (dynamic) interactions between proteins, technologies that allow detection and analysis of protein-protein interactions are essential for the study of cellular processes. Interaction data place a protein in its intracellular context, thus providing insights into its biological role. In addition, many of these interactions may represent interesting targets for drug development.

MAmmalian Protein-Protein Interaction Trap is a two-hybrid technology for detection of protein-protein interactions. While in the classical yeast two-hybrid technique a transcription factor is reconstituted upon interaction between two proteins, in MAPPIT the assay is based on functional complementation of a cytokine receptor signalling pathway. Read more »

ForwardMAPPIT allows the identification and detailed analysis of interactions between proteins. Modification-dependent protein interactions can be studied with the HeteromericMAPPIT variant. The ReverseMAPPIT mode can be used to screen for disrupters of protein-protein interactions including small organic molecules.  The three-hybrid mode, MASPIT - MAmmalian Small molecule-Protein Interaction Trap - enables the detection of interactions between proteins and other molecules including small organic molecules. KISS is an adaptation of MAPPIT that allows the detection of protein-protein interactions at their native subcellular location and is compatible with full size transmembrane proteins. Read more »

Analysis of designated interactions is done in binary assays using a luciferase-based reporter-assay or a STAT3 activation assay. To allow the identification of novel interactions, we developed a FACS-based assay to screen complex prey cDNA libraries. Alternatively, subsets of the human proteome can be rapidly screened in an array-based assay. Read more »

MAPPIT operates in a mammalian background, which brings about a number of interesting opportunities and advantages as compared to classical (yeast) two-hybrid methods or alternative approaches. Read more »

Since the publication of the original ForwardMAPPIT proof-of-concept paper in 2001, many new features have been added to the MAPPIT technology platform and the technology has been applied in diverse basic and applied research projects both in our own lab and those of many academic and industrial collaborators.

Virotrap is an additional protein-protein interaction detection method that was developed at the CRL. It also operates in mammalian cells but is based on a co-complex strategy. By capturing protein complexes in viral particles the cell lysis step is ommitted, thus reducing illegitimate interactions and facilitating the identification of weak interactions. For more information we refer to the paper Eyckerman et al. Nat. Commun. 7: 11416 (2016)