Project Summary The majority of excitatory synaptic transmission in the CNS synapses are mediated by the AMPA-type ionotropic glutamate receptor (AMPAR), a ligand gated ion channel activated by the neurotransmitter glutamate. The pore forming subunits of AMPARs (GluA1-4 subunits) assemble as homo- or hetero-tetramers. The native AMPARs co-assemble with a rich repertoire of transmembrane auxiliary subunits which belong to different protein families, such as TARP, GSG1L, CNIH, CKAMP, and synDIG. Incorporation of different auxiliary subunit confers AMPAR with unique ion channel gating kinetics, pharmacology, and in many cases trafficking regulation. The varying expression patters of auxiliary subunits in brain provides opportunities to produce drugs that target specific AMPAR auxiliary subunit combinations, which would have improved target specificity and less side effects over existing ones such as Perampanel (NAM) and ampakines (PAM) that bind to the common pore forming subunits of AMPARs (GluA1-4). Indeed, several NAMs that are selective to TARP gamma-8 containing AMPARs are already available and effective in seizures and pain. All the TARPs except for the TARP gamma-2 (encoded by CACNG2) are understudied and present in Phanos and IDG resource. Among these the biology is least understood for the type-II TARPs (CACNG5 and 7, which encodes TARP gamma-5 and 7), whose sequences are distant from the type-I TARPs (CACNG2, 3, 4, and 8, which encodes TARP gamma-2, 3, 4, and 8). Currently, it is established that type-II TARPs bind to calcium permeable AMPA-Rs (CP-AMPAR) that lacks the GluA2 subunit and regulate their functions. TARP gamma-5 is expressed in the CA2 of hippocampus and Bergmann glia of cerebellum, whereas TARP gamma-7 is enriched in cerebellar neurons (Purkinje, basket, stellate, granule, Bergmann glia, and Golgi cells). Consistently, CP-AMPARs were functionally detectable in these cell types. The function of CP-AMPARs is highly relevant to ischemic brain damages, brain tumors, addiction, fear-conditioning, and motor function. Compounds that selectively target type-II TARP containing CP-AMPARs are promising reagents for pharmacological manipulation to study the cellular function of these complexes and may facilitate the future development of drugs for treating the pathological conditions described above. We hypothesize that the type-II TARPs operate under different molecular mechanism from type-I TARPs, which do not specialize in CP-AMPAR modulation. To illuminate the structural basis for the mechanism of type-II TARP function, we propose to solve the high-resolution structures of a type-II TARP in complex with CP-AMPAR, which is currently missing. Based on the insights obtained from the structures of other AMPAR/auxiliary subunit complexes we predict that the differences in sidechain interaction network at the protein interaction interface determine the types of functional readout of modulation. The outcome of this research is expected to p...