The Molecular Mechanism Of GIT2and TRAF1Inhibits NF-κB Signaling Pathway

NF-κB transcription factors are critical regulators of immunity, stress response, apoptosis and play important roles in many biological processes, such as adaptative immunity and native immunity. TRAF(The TNF receptor-associated factor) family are scaffold proteins that mediate TNFR superfamily induced cell apoptosis. TRAF1was originally identified based on its ability to interact with the cytosolic domain of TNF receptor type2(TNFR2). TRAF1is composed of N terminal and C terminal domain. TRAF1is unique among TRAF proteins in that it lacks RING domain that found in the N-terminal regions of other TRAFs.GIT2(GPCR-kinase interacting proteins2) is ubiquitous multidomain proteins and is expressed in various tissue. It is identified as a scaffold regulator of various signal transducers, such as receptor internalization, cell migration and cell spreading. git2knockout mice show enlarged spleens, highly susceptible to infection which suggested that GIT2is important in immune response. However, whether GIT2directly regulates NF-κB signaling and detail mechanisms remain to be determined.We find that GIT2interacts with TRAF1, A20and NEMO etc using a yeast two-hybrid screen, which implicates that GIT2plays an important role in NF-κB related signaling pathways. We confirmed that GIT2interacts with TRAF1both in vivo and in vitro, and find the GIT2binding domain of TRAF1. Besides, we confirmed that GIT2and TRAF1co-inhibits NF-κB signaling pathway under the stimulation of TNF-a or IL-1β. GIT2inhibits NF-κB signaling pathway independent of its Arf-GAP activity. GIT2has a minor inhibition on the binding of TRAF1/TRAF2, but inhibits the binding of TRAF1/TRAF6. Besides, we demonstrated that GIT2and TRAF1co-regulate the ubiquitination of TRAF2and TRAF6.