The Rho Guanine Nucleotide Exchange Factor DRhoGEF2 Is a Genetic Modifier of the PI3K Pathway in Drosophila.
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ABSTRACT:
SUBMITTER: Chang YJ
PROVIDER: S-EPMC4807833 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundSmall GTPases of the Rho family are critical regulators of various cellular functions including actin cytoskeleton organization, activation of kinase cascades and mitogenesis. For this reason, a major objective has been to understand the mechanisms of Rho GTPase regulation. Here, we examine the function of a novel protein, Scambio, which shares homology with the DH-PH domains of several known guanine nucleotide exchange factors for Rho family members.ResultsScambio is located on human chromosome 14q11.1, encodes a protein of around 181 kDa, and is highly expressed in both heart and skeletal muscle. In contrast to most DH-PH-domain containing proteins, it binds the activated, GTP-bound forms of Rac and Cdc42. However, it fails to associate with V14RhoA. Immunofluorescence studies indicate that Scambio and activated Rac3 colocalize in membrane ruffles at the cell periphery. In accordance with these findings, Scambio does not activate either Rac or Cdc42 but rather, stimulates guanine nucleotide exchange on RhoA and its close relative, RhoC.ConclusionScambio associates with Rac in its activated conformation and functions as a guanine nucleotide exchange factor for Rho.
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Project description:Xpln is a guanine nucleotide-exchange factor (GEF) for Rho GTPases. A Dbl homology (DH) domain followed by a pleckstrin homology (PH) domain is a widely adopted GEF-domain architecture. The Xpln structure solely comprises these two domains. Xpln activates RhoA and RhoB, but not RhoC, although their GTPase sequences are highly conserved. The molecular mechanism of the selectivity of Xpln for Rho GTPases is still unclear. In this study, the crystal structure of the tandemly arranged DH-PH domains of mouse Xpln, with a single molecule in the asymmetric unit, was determined at 1.79?Å resolution by the multiwavelength anomalous dispersion method. The DH-PH domains of Xpln share high structural similarity with those from neuroepithelial cell-transforming gene 1 protein, PDZ-RhoGEF, leukaemia-associated RhoGEF and intersectins 1 and 2. The crystal structure indicated that the ?4-?5 loop in the DH domain is flexible and that the DH and PH domains interact with each other intramolecularly, thus suggesting that PH-domain rearrangement occurs upon RhoA binding.
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Project description:Collective cell migration plays crucial roles in tissue remodeling, wound healing, and cancer cell invasion. However, its underlying mechanism remains unknown. Previously, we showed that the RhoA-targeting guanine nucleotide exchange factor Solo (ARHGEF40) is required for tensile force-induced RhoA activation and proper organization of keratin-8/keratin-18 (K8/K18) networks. Here, we demonstrate that Solo knockdown significantly increases the rate at which Madin-Darby canine kidney cells collectively migrate on collagen gels. However, it has no apparent effect on the migratory speed of solitary cultured cells. Therefore, Solo decelerates collective cell migration. Moreover, Solo localized to the anteroposterior regions of cell-cell contact sites in collectively migrating cells and was required for the local accumulation of K8/K18 filaments in the forward areas of the cells. Partial Rho-associated protein kinase (ROCK) inhibition or K18 or plakoglobin knockdown also increased collective cell migration velocity. These results suggest that Solo acts as a brake for collective cell migration by generating pullback force at cell-cell contact sites via the RhoA-ROCK pathway. It may also promote the formation of desmosomal cell-cell junctions related to K8/K18 filaments and plakoglobin.
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Project description:G alpha 12/13-mediated pathways have been shown to be involved in various fundamental cellular functions in mammalian cells such as axonal guidance, apoptosis, and chemotaxis. Here, we identified a homologue of Rho-guanine nucleotide exchange factor (GEF) in Caenorhabditis elegans (CeRhoGEF), which functions downstream of gpa-12, the C. elegans homologue of G alpha 12/13. CeRhoGEF contains a PSD-95/Dlg/ZO-1 domain and a regulator of G protein signaling (RGS) domain upstream of the Dbl homology-pleckstrin homology region similar to mammalian RhoGEFs with RGS domains, PSD-95/Dlg/ZO-1-RhoGEF and leukemia-associated RhoGEF. It has been shown in mammalian cells that these RhoGEFs interact with activated forms of G alpha 12 or G alpha 13 through their RGS domains. We demonstrated by coimmunoprecipitation that the RGS domain of CeRhoGEF interacts with GPA-12 in an AIF4- activation-dependent manner and confirmed that the Dbl homology-pleckstrin homology domain of CeRhoGEF was capable of Rho-dependent signaling. These results proved conservation of the G alpha 12-RhoGEF pathway in C. elegans. Expression of DsRed or GFP under the control of the promoter of CeRhoGEF or gpa-12 revealed an overlap of their expression patterns in ventral cord motor neurons and several neurons in the head. RNA-mediated gene interference for CeRhoGEF and gpa-12 resulted in similar phenotypes such as embryonic lethality and sensory and locomotive defects in adults. Thus, the G alpha 12/13-RhoGEF pathway is likely to be involved in embryonic development and neuronal function in C. elegans.
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Project description:BACKGROUND:Intracranial aneurysm (IA) is usually a late-onset disease, affecting 1% to 3% of the general population and leading to life-threatening subarachnoid hemorrhage. Genetic susceptibility has been implicated in IAs, but the causative genes remain elusive. METHODS:We performed next-generation sequencing in a discovery cohort of 20 Chinese IA patients. Bioinformatics filters were exploited to search for candidate deleterious variants with rare and low allele frequency. We further examined the candidate variants in a multiethnic sample collection of 86 whole exome sequenced unsolved familial IA cases from 3 previously published studies. RESULTS:We identified that the low-frequency variant c.4394C>A_p.Ala1465Asp (rs2298808) of ARHGEF17 was significantly associated with IA in our Chinese discovery cohort (P=7.3×10-4; odds ratio=7.34). It was subsequently replicated in Japanese familial IA patients (P=0.039; odds ratio=4.00; 95% confidence interval=0.832-14.8) and was associated with IA in the large Chinese sample collection comprising 832 sporadic IA-affected and 599 control individuals (P=0.041; odds ratio=1.51; 95% confidence interval=1.02-Inf). When combining the sequencing data of all familial IA patients from 4 different ethnicities (ie, Chinese, Japanese, European American, and French-Canadian), we identified a significantly increased mutation burden for ARHGEF17 (21/106 versus 11/306; P=8.1×10-7; odds ratio=6.6; 95% confidence interval=2.9-15.8) in cases as compared with controls. In zebrafish, arhgef17 was highly expressed in the brain blood vessel. arhgef17 knockdown caused blood extravasation in the brain region. Endothelial lesions were identified exclusively on cerebral blood vessels in the arhgef17-deficient zebrafish. CONCLUSIONS:Our results provide compelling evidence that ARHGEF17 is a risk gene for IA.
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Project description:Nephrotic syndrome is a kidney disease featured by heavy proteinuria. It is caused by injury to the specialized epithelial cells called "podocytes" within the filtration unit of the kidney, glomerulus. Previous studies showed that hyperactivation of the RhoGTPase, Rac1, in podocytes causes podocyte injury and glomerulosclerosis (accumulation of extracellular matrix in the glomerulus). However, the mechanism by which Rac1 is activated during podocyte injury is unknown. Trio is a guanine nucleotide exchange factor (GEF) known to activate Rac1. By RNA-sequencing, we found that Trio mRNA is abundantly expressed in cultured human podocytes. Trio mRNA was also significantly upregulated in humans with minimal change disease and focal segmental glomerulosclerosis, two representative causes of nephrotic syndrome. Reduced expression of Trio in cultured human podocytes decreased basal Rac1 activity, cell size, attachment to laminin, and motility. Furthermore, while the pro-fibrotic cytokine, transforming growth factor ?1 increased Rac1 activity in control cells, it decreases Rac1 activity in cells with reduced Trio expression. This was likely due to simultaneous activation of the Rac1-GTPase activation protein, CdGAP. Thus, Trio is important in the basal functions of podocytes and may also contribute to glomerular pathology, such as sclerosis, via Rac1 activation.
| S-EPMC5855701 | biostudies-literature
Project description:Our study has shown that ARHGEF10 is a putative tumor suppressor in pancreatic ductal adenocarcinoma. To determine its functional mechanism, we perfomed microarray gene expression analysis of two pancreatic cancer cell line models of ARHGEF10 expression : MiaPACA2 cells in which ARHGEF10 was stably overexpressed, and Hs766T cells in which ARHGEF10 expression had been stably knocked down by short hairpin RNAs.
2019-09-05 | GSE131859 | GEO
Project description:Guanine nucleotide exchange factors (GEFs) play multiple functional roles in neurons. In a previous study, we reported that Arhgef4 (Rho guanine nucleotide exchange factor 4) functioned as a negative regulator of the excitatory synaptic function by sequestering postsynaptic density protein 95 (PSD-95). However, the role of Arhgef4 in behavior has not been examined. We performed comprehensive behavioral tests in knockout (KO) mice to investigate of the effects of Arhgef4 deficiency. We found that the expressed PSD-95 particle size was significantly increased in hippocampal neuronal cultures from Arhgef4 KO mice, which is consistent with the previous in vitro findings. Arhgef4 KO mice exhibited general motor activity and anxiety-like behavior comparable to those of the wild type littermates. However, spatial memory and object recognition memory were significantly enhanced in the Arhgef4 KO mice. Taken together, these data confirm the role of Arhgef4 as a negative synaptic regulator at the behavioral level.
| S-EPMC7649087 | biostudies-literature
Project description:Vertebrate-striated muscle is assumed to owe its remarkable order to the molecular ruler functions of the giant modular signaling proteins, titin and nebulin. It was believed that these two proteins represented unique results of protein evolution in vertebrate muscle. In this paper we report the identification of a third giant protein from vertebrate muscle, obscurin, encoded on chromosome 1q42. Obscurin is approximately 800 kD and is expressed specifically in skeletal and cardiac muscle. The complete cDNA sequence of obscurin reveals a modular architecture, consisting of >67 intracellular immunoglobulin (Ig)- or fibronectin-3-like domains with multiple splice variants. A large region of obscurin shows a modular architecture of tandem Ig domains reminiscent of the elastic region of titin. The COOH-terminal region of obscurin interacts via two specific Ig-like domains with the NH(2)-terminal Z-disk region of titin. Both proteins coassemble during myofibrillogenesis. During the progression of myofibrillogenesis, all obscurin epitopes become detectable at the M band. The presence of a calmodulin-binding IQ motif, and a Rho guanine nucleotide exchange factor domain in the COOH-terminal region suggest that obscurin is involved in Ca(2+)/calmodulin, as well as G protein-coupled signal transduction in the sarcomere.
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Project description:Epithelial tumor cells often acquire malignant properties, such as invasion/metastasis and uncontrolled cell growth, by undergoing epithelial-mesenchymal transition (EMT). However, the mechanisms by which EMT contributes to malignant progression remain elusive. Here we show that the Rho guanine nucleotide exchange factor (GEF) ARHGEF5 promotes tumor malignancy in a manner dependent on EMT status. We previously identified ARHGEF5, a member of the Dbl family of GEFs, as a multifunctional mediator of Src-induced cell invasion and tumor growth. In the present study, ARHGEF5 was upregulated during tumor growth factor-β-induced EMT in human epithelial MCF10A cells, and promoted cell migration by activating the Rho-ROCK pathway. ARHGEF5 was necessary for the invasive and in vivo metastatic activity of human colorectal cancer HCT116 cells. These findings underscore the crucial role of ARHGEF5 in cell migration and invasion/metastasis. An in vivo tumorigenesis assay revealed that ARHGEF5 had the potential to promote tumor growth via the phosphatidylinositol 3-kinase (PI3K) pathway. However, ARHGEF5 was not required for tumor growth in epithelial-like human colorectal cancer HCT116 and HT29 cells, whereas the growth of mesenchymal-like SW480 and SW620 cells depended on ARHGEF5. Induction of EMT by tumor necrosis factor-α or Slug in HCT116 cells resulted in the dependence of tumor growth on ARHGEF5. In these mesenchymal-like cells, Akt was activated via ARHGEF5 and its activity was required for tumor growth. Analysis of a transcriptome data set revealed that the combination of ARHGEF5 upregulation and E-cadherin downregulation or Snail upregulation was significantly correlated with poor prognosis in patients with colorectal cancers. Taken together, our findings suggest that EMT-induced ARHGEF5 activation contributes to the progression of tumor malignancy. ARHGEF5 may serve as a potential therapeutic target in a subset of malignant tumors that have undergone EMT.
| S-EPMC5047960 | biostudies-literature