Cytoplasmic retention sites in p190RhoGEF confer anti-apoptotic activity to an EGFP-tagged protein.
p190RhoGEF is a large multi-functional protein with guanine nucleotide exchange (GEF) activity. The C-terminal region of p190RhoGEF is a highly interactive domain that binds multiple factors, including proteins with anti-apoptotic activities. We now report that transfection of EGFP-tagged p190RhoGEF protects Neuro 2a cells from stress-induced apoptosis and that anti-apoptotic activity is localized to cytoplasmic retention sequences (CRS-1 and CRS-2) in the C-terminal region of p190RhoGEF. Cytoplasmic retention is conferred to an EGFP fluorescent marker when fused to either CRS-1 or CRS-2. Both cytoplasmic retention and anti-apoptotic activity are lost by deleting CRS-1 and CRS-2 in the p190RhoGEF sequence and can be recovered by restoring either CRS-1 or CRS-2 to the EGFP-tagged sequence. Since the CRS-1 and CRS-2 contain the JIP-1 and 14-3-3 binding sites, we propose that anti-apoptotic activity may be conferred by the binding of p190RhoGEF to JIP-1 or 14-3-3, possibly by altering their interactive properties or nucleocytoplasmic movements. Taken together, our findings support a model whereby multiple interactions of p190RhoGEF confer homeostatic properties to differentiated neurons and may link neuronal homeostasis to the regulation of NF-L expression.
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D001665:Binding Sites; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism; D002449:Cell Aggregation; D016923:Cell Death; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism*; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; D005982:Glutathione Transferase / Q000235:genetics; D049452:Green Fluorescent Proteins; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D020287:In Situ Nick-End Labeling / Q000379:methods; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins / Q000378:metabolism*; D018613:Microscopy, Confocal; D009687:Nuclear Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D011233:Precipitin Tests / Q000379:methods; D011485:Protein Binding; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012097:Repressor Proteins; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D020798:Two-Hybrid System Techniques
C513041:Arhgap35 protein, mouse; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; C121624:Mapkbp1 protein, mouse; D009687:Nuclear Proteins; D010446:Peptide Fragments; D016271:Proto-Oncogene Proteins c-myc; D012097:Repressor Proteins; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase