Annotated protein:	Rho guanine nucleotide exchange factor 7 (Beta-Pix) (PAK-interacting exchange factor beta). Gene symbol: ARHGEF7. Taxonomy: Rattus norvegicus (Rat). Uniprot ID: O55043
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SynGO gene info:	SynGO data @ ARHGEF7
Ontology domain:	Biological Process
SynGO term:	postsynaptic actin cytoskeleton organization (GO:0098974)
Synapse type(s):	hippocampus, GABAergic
Annotated paper:	Smith KR, et al. "GIT1 and betaPIX are essential for GABA(A) receptor synaptic stability and inhibitory neurotransmission" Cell Rep. 2014 Oct 9;9(1):298-310 PMID:25284783
Figure(s):	Fig.2, 3,
Annotation description:	Fig.2: "GIT1 Knockdown Alters GABAAR Surface Stability" - GIT1 regulates postsynaptic actin cytsoskeleton, thereby affecting staility of the postsynaptic specialization and GABAR localization. - "GIT1 knockdown neurons exhibited a significant decrease in surface GABAAR and VGAT cluster area compared to control (Figures 2A-2C), suggesting a possible role for GIT1 in maintaining the integrity of inhibitory pre- and postsynaptic domains in neuronal dendrites. Importantly, this effect could be rescued by coexpression of RNAi-resistant human GIT1 (hGIT1; Figures S2E and S2F). GIT1 knockdown also caused a large decrease in gephyrin cluster area (Figures 2D and 2E), suggesting that GIT1 is important for maintaining both GABAAR clusters and the gephyrin scaffold in neurons...Considering its role in other cell types, we hypothesized that GIT1 may be important for localizing F-actin regulatory pathways to inhibitory synapses. Therefore, we sought to determine whether surface GABAARs were sensitive to short-term disruption of the actin cytoskeleton by treating neurons with latrunculin-A, an inhibitor of actin polymerization (Renner et al., 2009). We found that a 30 min application of 3 μM latrunculin-A to neurons caused a significant decrease in surface GABAARs (Figures 2H and 2I), with no effect on extrasynaptic GABAAR populations (Figures S2G and S2H), suggesting that actin polymerization does indeed play an important role in the maintenance of inhibitory synapses. We then asked whether the inhibitory synapse effects we observed upon knockdown of GIT1 were due to altered F-actin regulation. We therefore treated control or GIT1 RNAi-expressing neurons with the F-actin-stabilizing drug, jasplakinolide (Hering et al., 2003), prior to surface biotinylation and western blot analysis (Figure 2J). As predicted, GIT1 RNAi caused a significant loss of surface GABAARs compared to control, which was restored by treatment with jasplakinolide (Figures 2J and 2K). This suggests that the decrease in surface GABAARs observed in GIT1-deficient neurons is caused by impaired F-actin regulation, and points toward a mechanism involving actin-regulatory proteins." Fig.3: "The Rac1 Activator βPIX Is Essential for GABAAR Surface Stability" - "GIT1's primary binding partner βPIX is one such actin-regulatory protein and is a strong candidate to collaborate with GIT1 in mediating actin regulation at inhibitory synapses. To test this hypothesis, we utilized RNAi to βPIX, which caused a significant reduction of βPIX expression levels (Figures S3A and S3B). βPIX knockdown in neurons caused a similar effect to that of GIT1 RNAi, reducing both surface GABAARs and VGAT cluster area (Figures 3A-3C). Surface biotinylation assays revealed the same phenotype, with βPIX knockdown neurons exhibiting reduced surface GABAAR levels compared with control cells (Figures 3D and 3E). We found GIT1 or βPIX knockdown had no effect on AMPA receptor clustering or extrasynaptic δ- containing GABAARs (Figures S3C-S3F), suggesting that this protein complex is important for synaptic GABAAR clustering only. We then sought to determine whether GIT1 and βPIX are important in controlling actin regulation at inhibitory synapses. We treated control neurons with phalloidin to label F-actin and found that ~80% of inhibitory synapses were positive for F-actin. GIT1 and βPIX knockdown caused a significant decrease in the percentage of inhibitory synapses containing F-actin (Figures 3F and 3G), suggesting that GIT1 and βPIX have an important role in controlling F-actin at inhibitory synapses. The effect of βPIX knockdown on surface GABAAR clusters was rescued by RNAi resistant human βPIX (hβPIX) showing the specificity of the RNAi. In contrast, versions of hβPIX that no longer have GEF activity for Rac1 (hβPIX-DN), or that contain a mutation of the βPIX SH3 domain (which is critical for coupling to a downstream effector, PAK [Hoelz et al., 2006]), were unable to rescue βPIX knockdown induced GABAAR declustering (Figures 3H and 3I). This suggests that the ability of βPIX to activate Rac1 and interact with PAK is essential for its role in maintaining inhibitory synapses and supports the idea that there exists a key actin regulatory mechanism controlling inhibitory synapse maintenance."
Evidence tracking, Biological System:	Cultured neurons
Evidence tracking, Protein Targeting:	RNAi / shRNA
Evidence tracking, Experiment Assay:	Confocal
Annotator(s):	Pim van Nierop (ORCID:0000-0003-0593-3443) Guus Smit (ORCID:0000-0002-2286-1587) Matthijs Verhage (ORCID:0000-0002-2514-0216)
Lab:	Department of Functional Genomics, Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Vrije Universiteit Amsterdam, 1081 HV Amsterdam, The Netherlands
SynGO annotation ID:	2790
Dataset release (version):	20231201
View annotation as GO-CAM model: