scholarly journals Regulation of type V adenylate cyclase by Ric8a, a guanine nucleotide exchange factor

2007 ◽  
Vol 406 (3) ◽  
pp. 383-388 ◽  
Author(s):  
Shyi-Chyi Wang ◽  
Hsing-Lin Lai ◽  
Yi-Ting Chiu ◽  
Ren Ou ◽  
Chuen-Lin Huang ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Guanine Nucleotide Exchange Factor ◽  
Dominant Negative ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
N Terminus ◽  
Type V

In the present study, we demonstrate that AC5 (type V adenylate cyclase) interacts with Ric8a through directly interacting at its N-terminus. Ric8a was shown to be a GEF (guanine nucleotide exchange factor) for several α subunits of heterotrimeric GTP binding proteins (Gα proteins) in vitro. Selective Gα targets of Ric8a have not yet been revealed in vivo. An interaction between AC5 and Ric8a was verified by pull-down assays, co-immunoprecipitation analyses, and co-localization in the brain. Expression of Ric8a selectively suppressed AC5 activity. Treating cells with pertussis toxin or expressing a dominant negative Gαi mutant abolished the suppressive effect of Ric8a, suggesting that interaction between the N-terminus of AC5 and a GEF (Ric8a) provides a novel pathway to fine-tune AC5 activity via a Gαi-mediated pathway.

scholarly journals Molecular Determinants of the Interaction between Coxsackievirus Protein 3A and Guanine Nucleotide Exchange Factor GBF1

2007 ◽  
Vol 81 (10) ◽  
pp. 5238-5245 ◽  
Author(s):  
Els Wessels ◽  
Daniël Duijsings ◽  
Kjerstin H. W. Lanke ◽  
Willem J. G. Melchers ◽  
Catherine L. Jackson ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Binding Domain ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Molecular Determinants ◽  
N Terminus ◽  
Insight Into

ABSTRACT The 3A protein of coxsackievirus B3 (CVB3), a small membrane protein that forms homodimers, inhibits endoplasmic reticulum-to-Golgi complex transport. Recently, we described the underlying mechanism by showing that the CVB3 3A protein binds to and inhibits the function of GBF1, a guanine nucleotide exchange factor for ADP-ribosylation factor 1 (Arf1), thereby interfering with Arf1-mediated COP-I recruitment. This study was undertaken to gain more insight into the molecular determinants underlying the interaction between 3A and GBF1. Here we show that 3A mutants that have lost the ability to dimerize are no longer able to bind to GBF1 and trap it on membranes. Moreover, we identify a conserved region in the N terminus of 3A that is crucial for GBF1 binding but not for 3A dimerization. Analysis of the binding domain in GBF1 showed that the extreme N terminus, the dimerization/cyclophilin binding domain, and the homology upstream of Sec7 domain are required for the interaction with 3A. In contrast to that of full-length GBF1, overexpression of a GBF1 mutant lacking its extreme N terminus failed to rescue the effects of 3A. Together, these data provide insight into the molecular requirements of the interaction between 3A and GBF1.

A Role for the Noncatalytic N Terminus in the Function of Cdc25, a Saccharomyces cerevisiae Ras-Guanine Nucleotide Exchange Factor

Genetics ◽  
2000 ◽  
Vol 154 (4) ◽  
pp. 1473-1484
Author(s):  
Reneé A Chen ◽  
Tamer Michaeli ◽  
Linda Van Aelst ◽  
Roymarie Ballester
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Heat Shock ◽  
Guanine Nucleotide Exchange Factor ◽  
Dominant Negative ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Ras Pathway ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
N Terminus

Abstract The Saccharomyces cerevisiae CDC25 gene encodes a guanine nucleotide exchange factor (GEF) for Ras proteins. Its catalytic domain is highly homologous to Ras-GEFs from all eukaryotes. Even though Cdc25 is the first Ras-GEF identified in any organism, we still know very little about how its function is regulated in yeast. In this work we provide evidence for the involvement of the N terminus of Cdc25 in the regulation of its activity. A truncated CDC25 lacking the noncatalytic C-terminal coding sequence was identified in a screen of high-copy suppressors of the heat-shock-sensitive phenotype of strains in which the Ras pathway is hyper-activated. The truncated gene acts as a dominant-negative mutant because it only suppresses the heat-shock sensitivity of strains that require the function of CDC25. Our two-hybrid assays and immunoprecipitation analyses show interactions between the N terminus of Cdc25 and itself, the C terminus, and the full-length protein. These results suggest that the dominant-negative effect may be a result of oligomerization with endogenous Cdc25. Further evidence of the role of the N terminus of Cdc25 in the regulation of its activity is provided by the mapping of the activating mutation of CDC25HS20 to the serine residue at position 365 in the noncatalytic N-terminal domain. This mutation induces a phenotype similar to activating mutants of other genes in the Ras pathway in yeast. Hence, the N terminus may exert a negative control on the catalytic activity of the protein. Taken together these results suggest that the N terminus plays a crucial role in regulating Cdc25 and consequently Ras activity, which in S. cerevisiae is essential for cell cycle progression.

scholarly journals Interaction of Ezrin with the Novel Guanine Nucleotide Exchange Factor PLEKHG6 Promotes RhoG-dependent Apical Cytoskeleton Rearrangements in Epithelial Cells

2007 ◽  
Vol 18 (12) ◽  
pp. 4780-4793 ◽  
Author(s):  
Romina D'Angelo ◽  
Sandra Aresta ◽  
Anne Blangy ◽  
Laurence Del Maestro ◽  
Daniel Louvard ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Morphological Changes ◽  
Guanine Nucleotide Exchange Factor ◽  
Dominant Negative ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Apical Pole ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

The mechanisms underlying functional interactions between ERM (ezrin, radixin, moesin) proteins and Rho GTPases are not well understood. Here we characterized the interaction between ezrin and a novel Rho guanine nucleotide exchange factor, PLEKHG6. We show that ezrin recruits PLEKHG6 to the apical pole of epithelial cells where PLEKHG6 induces the formation of microvilli and membrane ruffles. These morphological changes are inhibited by dominant negative forms of RhoG. Indeed, we found that PLEKHG6 activates RhoG and to a much lesser extent Rac1. In addition we show that ezrin forms a complex with PLEKHG6 and RhoG. Furthermore, we detected a ternary complex between ezrin, PLEKHG6, and the RhoG effector ELMO. We demonstrate that PLEKHG6 and ezrin are both required in macropinocytosis. After down-regulation of either PLEKHG6 or ezrin expression, we observed an inhibition of dextran uptake in EGF-stimulated A431 cells. Altogether, our data indicate that ezrin allows the local activation of RhoG at the apical pole of epithelial cells by recruiting upstream and downstream regulators of RhoG and that both PLEKHG6 and ezrin are required for efficient macropinocytosis.

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