Structural studies of TREM2-DAP12 signaling complexes

초록

Triggering Receptor Expressed on Myeloid cells 2 (TREM2) is a transmembrane protein specifically expressed in the microglia of the brain. TREM2 is an immune receptor that binds anionic ligands. Ligand binding induces phosphorylation of the immunoreceptor tyrosine-based activation motif (ITAM) within the intracellular domain of its adaptor protein DNAX activation protein 12 (DAP12), thereby activating downstream signaling pathways that regulate microglial survival, proliferation, phagocytosis, and motility. Among the risk factors of Alzheimer’s disease (AD) found to date, the TREM2 rare variants are one of the most dangerous factors for AD development. TREM2 signaling stimulate microglial Aβ plaque clearing in the early-stage AD. However, over-activated TREM2 signaling can induces in uncontrolled neuronal death in the late-stage AD. Although the extracellular domain structure was revealed by X-ray crystallography, the structure of full length TREM2 and DAP12 protein complex remain to be elucidated. We have successfully expressed TREM2 and DAP12 proteins in Hi5 insect cell infected with baculovirus, and purified human TREM2 and TREM2/DAP12 protein. Interestingly, When TREM2 was separated from Superdex 200 column using size-exclusion chromatography, TREM2 peak could be observed at about 13 mL, indicating that TREM2 should be assembled in a certain oligomeric state. Indeed, 2D classification of Cryo-EM imaging showed that TREM2 may form a dimeric architecture. However, the orientation of TREM2 particles observed in Cryo-EM imaging does not seem to be diverse, so to overcome this, we plan to increase the diversity of orientation by binding DAP12 and antibodies to TREM2 or making it in a lipid nanodisc state.