MAPPIT (mammalian protein-protein interaction trap) analysis of early steps in toll-like receptor signalling.
The mammalian protein-protein interaction trap (MAPPIT) is a two-hybrid technique founded on type I cytokine signal transduction. Thereby, bait and prey proteins are linked to signalling deficient cytokine receptor chimeras. Interaction of bait and prey and ligand stimulation restores functional JAK (Janus kinase)-STAT (signal transducers and activators of transcription) signalling, which ultimately leads to the transcription of a reporter or marker gene under the control of the STAT3-responsive rPAP1 promoter. In the subsequent protocol, we describe the use of MAPPIT to study early events in Toll-like receptor (TLR) signalling. We here demonstrate a "signalling interaction cascade" from TLR4 to IRAK-1.
Department of Medical Protein Research, Ghent University - VIB, Ghent, Belgium.
Peter
P
Ulrichts
Irma
I
Lemmens
Delphine
D
Lavens
Rudi
R
Beyaert
Jan
J
Tavernier
D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002460:Cell Line; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D011485:Protein Binding; D015398:Signal Transduction; D013997:Time Factors; D051193:Toll-Like Receptors / Q000032:analysis* / Q000235:genetics / Q000378:metabolism*; D019076:Transgenes / Q000235:genetics
D051193:Toll-Like Receptors