PKCα-mediated phosphorylation of the diacylglycerol kinase ζ MARCKS domain switches cell migration modes by regulating interactions with Rac1 and RhoA
| Autor: | Robin J. Parks, Elias Daher, Radoslav Zinoviev, Jean-Christian Maillet, Michael Phan, Ryan Ard, Stephen H. Gee |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Scaffold protein rac1 GTP-Binding Protein RHOA DGKζ diacylglycerol kinase ζ Biochemistry GST glutathione S-transferase Mice Cell Movement Ras homolog gene family PKCα protein kinase Cα PA phosphatidic acid member A (RhoA) Myristoylated Alanine-Rich C Kinase Substrate Mice Knockout biology diacylglycerol Chemistry protein kinase C (PKC) RhoGDI Rho guanine nucleotide dissociation inhibitor Cell biology phosphatidic acid Phosphorylation MARCKS myristoylated alanine-rich C kinase substrate Signal transduction diacylglycerol kinase (DGK DAGK) MHC myosin heavy chain Research Article Diacylglycerol Kinase Protein Kinase C-alpha PDZ domain SEM standard error of the mean Diglycerides 03 medical and health sciences Protein Domains Animals MARCKS Protein kinase A Molecular Biology Diacylglycerol kinase 030102 biochemistry & molecular biology Neuropeptides Ras-related C3 botulinum toxin substrate 1 (Rac1) Cell Biology CBB Coomassie brilliant blue Fibroblasts RBD Rho-binding domain MEF mouse embryonic fibroblast GDI guanine nucleotide dissociation inhibitor syntrophin 030104 developmental biology scaffold protein Dystrophin-Associated Proteins biology.protein rhoA GTP-Binding Protein PBD p21-binding domain SD standard deviation DAG diacylglycerol HA hemagglutinin |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X |
| Popis: | Cells can switch between Rac1 (lamellipodia-based) and RhoA (blebbing-based) migration modes, but the molecular mechanisms regulating this shift are not fully understood. Diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid, forms independent complexes with Rac1 and RhoA, selectively dissociating each from their common inhibitor RhoGDI. DGKζ catalytic activity is required for Rac1 dissociation but is dispensable for RhoA dissociation; instead, DGKζ stimulates RhoA release via a kinase-independent scaffolding mechanism. The molecular determinants that mediate the selective targeting of DGKζ to Rac1 or RhoA signaling complexes are unknown. Here, we show that protein kinase Cα (PKCα)-mediated phosphorylation of the DGKζ MARCKS domain increased DGKζ association with RhoA and decreased its interaction with Rac1. The same modification also enhanced DGKζ interaction with the scaffold protein syntrophin. Expression of a phosphomimetic DGKζ mutant stimulated membrane blebbing in mouse embryonic fibroblasts and C2C12 myoblasts, which was augmented by inhibition of endogenous Rac1. DGKζ expression in differentiated C2 myotubes, which have low endogenous Rac1 levels, also induced substantial membrane blebbing via the RhoA-ROCK pathway. These events were independent of DGKζ catalytic activity, but dependent upon a functional C-terminal PDZ-binding motif. Rescue of RhoA activity in DGKζ-null cells also required the PDZ-binding motif, suggesting that syntrophin interaction is necessary for optimal RhoA activation. Collectively, our results define a switch-like mechanism whereby DGKζ phosphorylation by PKCα plays a role in the interconversion between Rac1 and RhoA signaling pathways that underlie different cellular migration modes. |
| Databáze: | OpenAIRE |
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