Type A GABA receptors (GABAA receptors) are the major inhibitory neurotransmitter receptors in the central nervous system. They can response to the neurotransmitter γ-aminobutyric acid (GABA) and are permeable to chloride ions, thus can inhibit neural excitability. The localization of GABAA receptors on the nerve cell membrane has an important role in the regulation of neural signal production and transmission. Numerous studies have shown that dysregulation of GABAA receptor distribution at the cell membrane level is closely associated with many neurological disorders, including a variety of neurodevelopmental disorders, such as schizophrenia, bipolar disorder, and autism; and chronic neurological disorders, such as anxiety, depression, and insomnia. In addition, GABAA receptors are important drug targets for neurological disorders. Therefore, the important role of dynamic regulation of GABAA receptors for the nervous system is unquestionable and has always been the research focus. However, there are still a number of questions about the molecular mechanism of GABAA receptor transport regulation on cell membranes remained to be answered.
On 12 Jan 2021, the research team led by Jianchao Li from School of Medicine SCUT, in collaboration with the research teams led by Prof. Chao Wang and Prof. Wei Xiong from School of Life Science USTC, published a research article entitle “Structural basis of GABARAP-mediated GABAA receptor trafficking and functions on GABAergic synaptic transmission” in the journal Nature Communications. They used a combination approach of biochemistry, structural biology, molecular neurology, electrophysiology, and chemical biology, to reveal the molecular mechanism of GABARAP-mediated GABAA receptor trafficking.
GABARAP was first identified as a protein that interacts with the γ2 subunit of GABAA receptor and is one of the important proteins involved in the process of GABAA receptor cell membrane localization. Previous studies have shown that GABARAP can increase the distribution of GABAA receptors in COS-7 cell membranes and hippocampal neuronal cell membranes, but its specific pathway of action remains unclear. In this study, the researchers found that an 18-amino acid short peptide (γ2-GIM) from the TM3-TM4 intracellular segment of the γ2 subunit directly binds GABARAP through in vitro biochemical assays. The team further discovered the specificity of γ2-GIM in binding GABARAP and GABARAPL1, and resolved the three-dimensional structure of the GABARAPL1/γ2-GIM complex, and determined the molecular mechanism of complex assembly.
Next, by co-expressing GABAA receptor and GABARAP in HEK293 cells, the researchers found that GABARAP significantly increased the GABA-mediated chloride current by electrophysiological means. This effect could be further enhanced by the endocytosis inhibitor Dynasore. In contrast, the GABA-mediated chloride current was significantly reduced when the exocytosis inhibitor Brefeldin A was added, demonstrating that GABARAP acts in a pathway that facilitates the receptor exocytosis. Finally, using an Ankyrin-derived interfering peptide previously developed by the same group (Nat Chem Biol, 2018; PNAS, 2020), the researchers found that disrupting the interaction between GABAA receptor and GABARAP in mouse brain caused a significant reduction of the miniature inhibitory postsynaptic currents (mIPSCs) in motor cortex neurons.
This work used various research methods to study the structure and function of the GABAA receptor complex with GABARAP, and revealed the molecular mechanism of how GABARAP regulates the dynamic distribution level of GABAA receptors on cell membranes. This work can serve as the foundation for further development of potent and specific drugs to treat related neurological disorders.
Prof. Jianchao Li from South China University of Technology, Prof. Chao Wang and Prof. Wei Xiong from University of Science and Technology China are listed as the co-corresponding authors of the research paper.
