Abstract
We have previously shown that the glutamate receptor interacting protein 1 (GRIP1) splice forms GRIP1a/b and GRIP1c4-7 are present at the GABAergic postsynaptic complex. Nevertheless the role that these GRIP1 protein isoforms play at the GABAergic postsynaptic complex is not known. We are now showing that GRIP1c4-7 and GRIP1a/b interact with gephyrin, the main postsynaptic scaffold protein of GABAergic and glycinergic synapses. Gephyrin coprecipitates with GRIP1c4-7 or GRIP1a/b from rat brain extracts and from extracts of HEK293 cells that have been cotransfected with gephyrin and one of the GRIP1 protein isoforms. Moreover, purified gephyrin binds to purified GRIP1c4-7 or GRIP1a/b, indicating that gephyrin directly interacts with the common region of these GRIP1 proteins, which includes PDZ domains 4–7. An engineered deletion construct of GRIP1a/b (GRIP1a4–7), which both contains the aforementioned common region and binds to gephyrin, targets to the postsynaptic GABAergic complex of transfected cultured hippocampal neurons. In these hippocampal cultures, endogenous gephyrin colocalizes with endogenous GRIP1c4-7 and GRIP1a/b in over 90% of the GABAergic synapses. Double-labeling electron microscopy immunogold reveals that in the rat brain GRIP1c4-7 and GRIP1a/b colocalize with gephyrin at the postsynaptic complex of individual synapses. These results indicate that GRIP1c4-7 and GRIP1a/b colocalize and interact with gephyrin at the GABAergic postsynaptic complex and suggest that this interaction plays a role in GABAergic synaptic function.
