scholarly journals Oligomerization of the Sec7 domain Arf guanine nucleotide exchange factor GBF1 is dispensable for Golgi localization and function but regulates degradation

2016 ◽  
Vol 310 (6) ◽  
pp. C456-C469 ◽  
Author(s):  
Jay M. Bhatt ◽  
Ekaterina G. Viktorova ◽  
Theodore Busby ◽  
Paulina Wyrozumska ◽  
Laura E. Newman ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Sec7 Domain ◽  
And Function ◽  
In Cells

Members of the large Sec7 domain-containing Arf guanine nucleotide exchange factor (GEF) family have been shown to dimerize through their NH2-terminal dimerization and cyclophilin binding (DCB) and homology upstream of Sec7 (HUS) domains. However, the importance of dimerization in GEF localization and function has not been assessed. We generated a GBF1 mutant (91/130) in which two residues required for oligomerization (K91 and E130 within the DCB domain) were replaced with A and assessed the effects of these mutations on GBF1 localization and cellular functions. We show that 91/130 is compromised in oligomerization but that it targets to the Golgi in a manner indistinguishable from wild-type GBF1 and that it rapidly exchanges between the cytosolic and membrane-bound pools. The 91/130 mutant appears active as it integrates within the functional network at the Golgi, supports Arf activation and COPI recruitment, and sustains Golgi homeostasis and cargo secretion when provided as a sole copy of functional GBF1 in cells. In addition, like wild-type GBF1, the 91/130 mutant supports poliovirus RNA replication, a process requiring GBF1 but believed to be independent of GBF1 catalytic activity. However, oligomerization appears to stabilize GBF1 in cells, and the 91/130 mutant is degraded faster than the wild-type GBF1. Our data support a model in which oligomerization is not a key regulator of GBF1 activity but impacts its function by regulating the cellular levels of GBF1.

scholarly journals Mutations in the GCD7 subunit of yeast guanine nucleotide exchange factor eIF-2B overcome the inhibitory effects of phosphorylated eIF-2 on translation initiation.

1994 ◽  
Vol 14 (5) ◽  
pp. 3208-3222 ◽  
Author(s):  
C R Vazquez de Aldana ◽  
A G Hinnebusch
Keyword(s):  
Translation Initiation ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Inhibitory Effects ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Suppressor Mutations ◽  
Nucleotide Exchange Factor ◽  
In Cells

Phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2 (eIF-2 alpha) impairs translation initiation by inhibiting the guanine nucleotide exchange factor for eIF-2, known as eIF-2B. In Saccharomyces cerevisiae, phosphorylation of eIF-2 alpha by the protein kinase GCN2 specifically stimulates translation of GCN4 mRNA in addition to reducing general protein synthesis. We isolated mutations in several unlinked genes that suppress the growth-inhibitory effect of eIF-2 alpha phosphorylation catalyzed by mutationally activated forms of GCN2. These suppressor mutations, affecting eIF-2 alpha and the essential subunits of eIF-2B encoded by GCD7 and GCD2, do not reduce the level of eIF-2 alpha phosphorylation in cells expressing the activated GCN2c kinase. Four GCD7 suppressors were shown to reduce the derepression of GCN4 translation in cells containing wild-type GCN2 under starvation conditions or in GCN2c strains. A fifth GCD7 allele, constructed in vitro by combining two of the GCD7 suppressors mutations, completely impaired the derepression of GCN4 translation, a phenotype characteristic of deletions in GCN1, GCN2, or GCN3. This double GCD7 mutation also completely suppressed the lethal effect of expressing the mammalian eIF-2 alpha kinase dsRNA-PK in yeast cells, showing that the translational machinery had been rendered completely insensitive to phosphorylated eIF-2. None of the GCD7 mutations had any detrimental effect on cell growth under nonstarvation conditions, suggesting that recycling of eIF-2 occurs efficiently in the suppressor strains. We propose that GCD7 and GCD2 play important roles in the regulatory interaction between eIF-2 and eIF-2B and that the suppressor mutations we isolated in these genes decrease the susceptibility of eIF-2B to the inhibitory effects of phosphorylated eIF-2 without impairing the essential catalytic function of eIF-2B in translation initiation.

scholarly journals REI-1 Is a Guanine Nucleotide Exchange Factor Regulating RAB-11 Localization and Function in C. elegans Embryos

2015 ◽  
Vol 35 (2) ◽  
pp. 211-221 ◽  
Author(s):  
Aisa Sakaguchi ◽  
Miyuki Sato ◽  
Katsuya Sato ◽  
Keiko Gengyo-Ando ◽  
Tomohiro Yorimitsu ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
C Elegans ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
And Function

scholarly journals Multifaceted Roles of the Ras Guanine-Nucleotide Exchange Factor ChRgf in Development, Pathogenesis, and Stress Responses of Colletotrichum higginsianum

2017 ◽  
Vol 107 (4) ◽  
pp. 433-443 ◽  
Author(s):  
Qiongnan Gu ◽  
Meijuan Chen ◽  
Junbin Huang ◽  
Yangdou Wei ◽  
Tom Hsiang ◽  
...  
Keyword(s):  
Stress Responses ◽  
Vegetative Growth ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Colletotrichum Higginsianum ◽  
Nucleotide Exchange Factor

The infection process of Colletotrichum higginsianum, which causes a disease of crucifers, involves several key steps: conidial germination, appressorial formation, appressorial penetration, and invasive growth in host tissues. In this study, the ChRgf gene encoding a Ras guanine-nucleotide exchange factor protein was identified by screening T-DNA insertion mutants generated from Agrobacterium tumefaciens-mediated transformation that were unable to cause disease on the host Arabidopsis thaliana. Targeted gene deletion of ChRgf resulted in a null mutant (ΔChrgf-42) with defects in vegetative growth, hyphal morphology, and conidiation, and poor surface attachment and low germination on hydrophobic surfaces; however, there were no apparent differences in appressorial turgor pressure between the wild type and the mutant. The conidia of the mutant were unable to geminate on attached Arabidopsis leaves and did not cause any disease symptoms. Intracellular cyclic adenosine monophosphate levels in the ΔChrgf mutant were lower than that of the wild type. Our results suggest that ChRgf is a key regulator in response to salt and osmotic stresses in C. higginsianum, and indicate that it is involved in fungal pathogenicity. This gene seems to act as an important modulator upstream of several distinct signaling pathways that are involved in regulating vegetative growth, conidiation, infection-related structure development, and stress responses of C. higginsianum.

Mutations in the GCD7 subunit of yeast guanine nucleotide exchange factor eIF-2B overcome the inhibitory effects of phosphorylated eIF-2 on translation initiation

1994 ◽  
Vol 14 (5) ◽  
pp. 3208-3222
Author(s):  
C R Vazquez de Aldana ◽  
A G Hinnebusch
Keyword(s):  
Translation Initiation ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Inhibitory Effects ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Suppressor Mutations ◽  
Nucleotide Exchange Factor ◽  
In Cells

Phosphorylation of the alpha subunit of eukaryotic translation initiation factor 2 (eIF-2 alpha) impairs translation initiation by inhibiting the guanine nucleotide exchange factor for eIF-2, known as eIF-2B. In Saccharomyces cerevisiae, phosphorylation of eIF-2 alpha by the protein kinase GCN2 specifically stimulates translation of GCN4 mRNA in addition to reducing general protein synthesis. We isolated mutations in several unlinked genes that suppress the growth-inhibitory effect of eIF-2 alpha phosphorylation catalyzed by mutationally activated forms of GCN2. These suppressor mutations, affecting eIF-2 alpha and the essential subunits of eIF-2B encoded by GCD7 and GCD2, do not reduce the level of eIF-2 alpha phosphorylation in cells expressing the activated GCN2c kinase. Four GCD7 suppressors were shown to reduce the derepression of GCN4 translation in cells containing wild-type GCN2 under starvation conditions or in GCN2c strains. A fifth GCD7 allele, constructed in vitro by combining two of the GCD7 suppressors mutations, completely impaired the derepression of GCN4 translation, a phenotype characteristic of deletions in GCN1, GCN2, or GCN3. This double GCD7 mutation also completely suppressed the lethal effect of expressing the mammalian eIF-2 alpha kinase dsRNA-PK in yeast cells, showing that the translational machinery had been rendered completely insensitive to phosphorylated eIF-2. None of the GCD7 mutations had any detrimental effect on cell growth under nonstarvation conditions, suggesting that recycling of eIF-2 occurs efficiently in the suppressor strains. We propose that GCD7 and GCD2 play important roles in the regulatory interaction between eIF-2 and eIF-2B and that the suppressor mutations we isolated in these genes decrease the susceptibility of eIF-2B to the inhibitory effects of phosphorylated eIF-2 without impairing the essential catalytic function of eIF-2B in translation initiation.

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