GFP‐p65 knock‐in mice as a tool to study NF‐κB dynamics in vivo

Abstract The nuclear factor κB (NF‐κB) signaling pathway regulates immune and inflammatory responses and is implicated in the pathogenesis of multiple diseases. The principal mechanism controlling NF‐κB activation depends on the association of NF‐κB transcription factor dimers with IκB molecules, which prevents the accumulation of NF‐κB in the nucleus and the activation of target gene transcription. Monitoring the nucleocytoplalsmic shuttling of NF‐κB factors is a reliable method to study the dynamic regulation of NF‐κB activity. Here, we generated knock‐in mice expressing a fusion protein between the green fluorescent protein (GFP) and the p65/RelA NF‐κB subunit (GFP‐p65) from the endogenous p65 genomic locus. Homozygous GFP‐p65 mice developed normally and showed normal NF‐κB activation, demonstrating that the GFP‐p65 fusion protein functionally substitutes for wild‐type p65. Live imaging of primary cells from GFP‐p65 mice allowed real‐time monitoring of p65 nucleocytoplasmic shuttling upon NF‐κB activation. Therefore, the GFP‐p65 knock‐in mice constitute an invaluable tool for studying the dynamic regulation of NF‐κB. genesis 47:323–329, 2009. © 2009 Wiley‐Liss, Inc.