Detection of recombinant and cellular MALT1 paracaspase activity.
MALT1 (mucosa-associated lymphoid tissue protein 1) is a key regulator of antigen-induced NF-κB activation in the adaptive immune response. Activation of proteolytic activity of the MALT1 paracaspase was shown to boost the immune response. Additionally, MALT1 proteolytic activity is essential for the survival of MALT1-dependent lymphoma, such as the activated B-cell type (ABC) of diffuse large B-cell lymphoma (DLBCL) or MALT lymphoma. The functional impact of MALT1 paracaspase on T-cell activation and lymphomagenesis suggests that MALT1 is a promising therapeutic target for the treatment of autoimmune diseases and distinct lymphoma entities. To evaluate the requirement of MALT1 in further detail, direct measurement of its activity status is of great importance. We have established a fluorogenic cleavage assay which can be used to measure activity of recombinant and cellular MALT1. Here we describe the basis of the cleavage assay and include a detailed protocol for recombinant production of MALT1 and also the cellular immunoprecipitation of endogenous MALT1 to determine its proteolytic activity.
Research Unit Cellular Signal Integration, Institute of Molecular Toxicology and Pharmacology, Helmholtz Zentrum München, Ingolstädter Landstr. 1, 85764, Neuherberg, Germany.
Daniel
D
Nagel
Daniel
D
Krappmann
D020169:Caspases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation; D057075:Enzyme Assays; D015870:Gene Expression; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D000074884:Mucosa-Associated Lymphoid Tissue Lymphoma Translocation 1 Protein; D009363:Neoplasm Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D059748:Proteolysis; D011994:Recombinant Proteins
D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D020169:Caspases; C400109:MALT1 protein, human; D000074884:Mucosa-Associated Lymphoid Tissue Lymphoma Translocation 1 Protein