scholarly journals Protein Kinase A (PKA) Type I Interacts with P-Rex1, a Rac Guanine Nucleotide Exchange Factor

2016 ◽  
Vol 291 (12) ◽  
pp. 6182-6199 ◽  
Author(s):  
Lydia Chávez-Vargas ◽  
Sendi Rafael Adame-García ◽  
Rodolfo Daniel Cervantes-Villagrana ◽  
Alejandro Castillo-Kauil ◽  
Jessica G. H. Bruystens ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Guanine Nucleotide Exchange Factor ◽  
Type I ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Kinase A

scholarly journals Protein Kinase A-Dependent Phosphorylation Modulates β1Pix Guanine Nucleotide Exchange Factor Activity through 14-3-3β Binding

2007 ◽  
Vol 28 (5) ◽  
pp. 1679-1687 ◽  
Author(s):  
Ahmed Chahdi ◽  
Andrey Sorokin
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpases ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Kinase A

ABSTRACT β1Pix is a guanine nucleotide exchange factor (GEF) for the small GTPases Rac and Cdc42 which has been shown to mediate signaling pathways leading to cytoskeletal reorganization. In the present study, we show that the basal association between endogenous βPix and endogenous 14-3-3β was increased after forskolin stimulation and significantly inhibited by protein kinase A inhibitor. However, forskolin stimulation failed to increase the interaction between 14-3-3β and a β1Pix mutant that is insensitive to protein kinase A phosphorylation, β1Pix(S516A, T526A). We present evidence indicating that forskolin-induced binding of 14-3-3β to β1Pix results in inhibition of Rac1 GTP loading in 293 cells and in vitro. Furthermore, we show that deletion of 10 amino acid residues within the leucine zipper domain is sufficient to block β1Pix homodimerization and 14-3-3β binding and modulates β1Pix-GEF activity. These residues also play a crucial role in β1Pix intracellular localization. These results indicate that 14-3-3β negatively affects the GEF activity of dimeric β1Pix only. Altogether, these results provide a mechanistic insight into the role of 14-3-3β in modulating β1Pix-GEF activity.

Phosphorylation of the guanine-nucleotide-exchange factor CalDAG-GEFI by protein kinase A regulates Ca2+-dependent activation of platelet Rap1b GTPase

2013 ◽  
Vol 453 (1) ◽  
pp. 115-123 ◽  
Author(s):  
Gianni Francesco Guidetti ◽  
Daria Manganaro ◽  
Alessandra Consonni ◽  
Ilaria Canobbio ◽  
Cesare Balduini ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
293 Cells ◽  
Kinase A

In blood platelets the small GTPase Rap1b is activated by cytosolic Ca2+ and promotes integrin αIIbβ3 inside-out activation and platelet aggregation. cAMP is the major inhibitor of platelet function and antagonizes Rap1b stimulation through a mechanism that remains unclear. In the present study we demonstrate that the Ca2+-dependent exchange factor for Rap1b, CalDAG-GEFI (calcium and diacylglycerol-regulated guanine-nucleotide-exchange factor I), is a novel substrate for the cAMP-activated PKA (protein kinase A). CalDAG-GEFI phosphorylation occurred in intact platelets treated with the cAMP-increasing agent forskolin and was inhibited by the PKA inhibitor H89. Purified recombinant CalDAG-GEFI was also phosphorylated in vitro by the PKA catalytic subunit. By screening a panel of specific serine to alanine residue mutants, we identified Ser116 and Ser586 as PKA phosphorylation sites in CalDAG-GEFI. In transfected HEK (human embryonic kidney)-293 cells, as well as in platelets, forskolin-induced phosphorylation of CalDAG-GEFI prevented the activation of Rap1b induced by the Ca2+ ionophore A23187. In platelets this effect was associated with the inhibition of aggregation. Moreover, cAMP-mediated inhibition of Rap1b was lost in HEK-293 cells transfected with a double mutant of CalDAG-GEFI unable to be phosphorylated by PKA. The results of the present study demonstrate that phosphorylation of CalDAG-GEFI by PKA affects its activity and represents a novel mechanism for cAMP-mediated inhibition of Rap1b in platelets.

scholarly journals Modulation of Rho Guanine Exchange Factor Lfc Activity by Protein Kinase A-Mediated Phosphorylation

2009 ◽  
Vol 29 (21) ◽  
pp. 5963-5973 ◽  
Author(s):  
David Meiri ◽  
Melissa A. Greeve ◽  
Andrea Brunet ◽  
Dina Finan ◽  
Clark D. Wells ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Dependent Manner ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Dynein Motor ◽  
Nucleotide Exchange Factor ◽  
Kinase A ◽  
Exchange Activity

ABSTRACT Lfc is a guanine nucleotide exchange factor (GEF) for Rho that demonstrates an unusual ability to associate with microtubules. While several phosphorylated residues have been detected in the Lfc polypeptide, the mechanism(s) by which phosphorylation regulates the exchange activity of Lfc remains unclear. We confirm that Lfc is a phosphorylated protein and demonstrate that 14-3-3 interacts directly and in a phosphorylation-dependent manner with Lfc. We identify AKAP121 as an Lfc-binding protein and show that Lfc is phosphorylated in an AKAP-dependent manner by protein kinase A (PKA). Forskolin treatment induced 14-3-3 binding to Lfc and suppressed the exchange activity of wild-type Lfc on RhoA. Importantly, a mutant of Lfc that is unable to associate with 14-3-3 proteins was resistant to inhibition by forskolin. Tctex-1, a dynein motor light chain, binds to Lfc in a competitive manner with 14-3-3.

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