Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The dbl oncogene encodes a prototype member of the Rho GTPase guanine nucleotide exchange factor (GEF) family. Oncogenic activation of proto-Dbl occurs through truncation of the N-terminal 497 residues. The C-terminal half of proto-Dbl includes residues 498 to 680 and 710 to 815, which fold into the Dbl homology (DH) domain and the pleckstrin homology (PH) domain, respectively, both of which are essential for cell transformation via the Rho GEF activity or cytoskeletal targeting function. Here we have investigated the mechanism of the apparent negative regulation of proto-Dbl imposed by the N-terminal sequences. Deletion of the N-terminal 285 or C-terminal 100 residues of proto-Dbl did not significantly affect either its transforming activity or GEF activity, while removal of the N-terminal 348 amino acids resulted in a significant increase in both transformation and GEF potential. Proto-Dbl displayed a mostly perinuclear distribution pattern, similar to a polypeptide derived from its N-terminal sequences, whereas onco-Dbl colocalized with actin stress fibers, like the PH domain. Coexpression of the N-terminal 482 residues with onco-Dbl resulted in disruption of its cytoskeletal localization and led to inhibition of onco-Dbl transforming activity. The apparent interference with the DH and PH functions by the N-terminal sequences can be rationalized by the observation that the N-terminal 482 residues or a fragment containing residues 286 to 482 binds specifically to the PH domain, limiting the access of Rho GTPases to the catalytic DH domain and masking the intracellular targeting function of the PH domain. Taken together, our findings unveiled an autoinhibitory mode of regulation of proto-Dbl that is mediated by the intramolecular interaction between its N-terminal sequences and PH domain, directly impacting both the GEF function and intracellular distribution.
Mol Cell Biol 2001 Mar
PMID:Autoinhibition mechanism of proto-Dbl. 1123 83

Ras is a small GTPase that is activated by upstream guanine nucleotide exchange factors, one of which is Ras-GRF2. GRF2 is a widely expressed protein with several recognizable sequence motifs, including a Ras exchanger motif (REM), a PEST region containing a destruction box (DB), and a Cdc25 domain. The Cdc25 domain possesses guanine nucleotide exchange factor activity and interacts with Ras. Herein we examine if the DB motif in GRF2 results in proteolysis via the ubiquitin pathway. Based on the solved structure of the REM and Cdc25 regions of the Son-of-sevenless (Sos) protein, the REM may stabilize the Cdc25 domain during Ras binding. The DB motif of GRF2 is situated between the REM and the Cdc25 domains, tempting speculation that it may be exposed to ubiquitination machinery upon Ras binding. GRF2 protein levels decrease dramatically upon activation of GRF2, and dominant-negative Ras induces degradation of GRF2, demonstrating that signaling downstream of Ras is not required for the destruction of GRF2 and that binding to Ras is important for degradation. GRF2 is ubiquitinated in vivo, and this can be detected using mass spectrometry. In the presence of proteasome inhibitors, Ras-GRF2 accumulates as a high-molecular-weight conjugate, suggesting that GRF2 is destroyed by the 26S proteasome. Deleting the DB reduces the ubiquitination of GRF2. GRF2 lacking the Cdc25 domain is not ubiquitinated, suggesting that a protein that cannot bind Ras cannot be properly targeted for destruction. Point mutations within the Cdc25 domain that eliminate Ras binding also eliminate ubiquitination, demonstrating that binding to Ras is necessary for ubiquitination of GRF2. We conclude that conformational changes induced by GTPase binding expose the DB and thereby target GRF2 for destruction.
Mol Cell Biol 2001 Mar
PMID:Ras binding triggers ubiquitination of the Ras exchange factor Ras-GRF2. 1123 45

BetaPix, a Pak-interacting guanine nucleotide exchange factor is known to be involved in the regulation of Cdc42/Rac GTPases and Pak kinase activity. Currently, three 1Pix isoforms, betaPix-a, -b, and -c have been reported. In this study, the cDNA of a novel Pix splice variant was isolated from a mouse brain cDNA library. The cloned betaPix isoform, named betaPix-d, lacks leucine zipper domain that is present in other Pix isoforms, and has a 11 amino acid addition at carboxyl terminus and distinct 3'-UTR Analysis of the tissue distribution of betaPix-d using RT-PCR revealed that its message was present mainly in brain and testis but in lower levels in heart, spleen, lung, liver, skeletal muscle and kidney. In situ hybridization studies with the 13Pix-d specific probes in the rat embryo show that betaPix-d isoform is expressed mainly in the central nervous system. Moreover, temporal expression pattern of the isoform is correlated with the active neurogenesis period in the cerebral cortex and cerebellum during rat brain development. These findings suggest that betaPix-d isoform may be developmentally regulated.
Mol Cells 2001 Feb 28
PMID:Molecular cloning and characterization of a novel mouse betaPix isoform. 1126 27

RalA GTPase, a member of Ras superfamily proteins, shows alternative forms between the active GTP-binding and the inactive GDP-binding states. Ral-specific guanine nucleotide exchange factor such as RalGDS interacts with activated Ras and cooperates with Ras indicating that Ral can be activated through Ras signaling pathway. Another activation path for Ral are through Ca2+-dependent but Ras-independent manner. In this study, studies were carried out to examine possible effects of Ca2+ and calmodulin, Ca2+-binding protein, directly on the GTP/GDP-binding state to recombinant unprenylated GST-RalA proteins. The results showed that Ca2+ stimulated the binding of GTP to RalA, whereas it reduced the binding of GDP to RalA. However, it does not involve a high affinity association of Ca2+ with RalA. Ca2+/calmodulin stimulated the GTPase activity of RalA. These results indicate that Ca2+ alone activates RalA by stimulating GTP-binding to RalA and Ca2+/calmodulin inactivates RalA by increasing the activity of RalGTPase.
Exp Mol Med 2001 Mar 31
PMID:Regulation of GTP-binding state in RalA through Ca2+ and calmodulin. 1132 87

Stimulation of the PRL receptor (PRLr) results in the activation of the guanine nucleotide exchange factor (GEF) p95Vav1 with corresponding alterations in cytoarchitecture and cell motility. To better understand the mechanisms involved in the regulation of Vav1 activity, the role of the tyrosine kinase p70Tec was examined. Coimmunoprecipitation and in vitro kinase assays revealed that ligand stimulation of the PRLr resulted in the rapid activation of Tec and its concomitant association with the PRLR: When coexpressed in COS-1 cells, both Vav1 and Tec were found to associate with the PRLr in the presence of ligand. In the absence of receptor, a constitutive complex between Vav1 and Tec was noted. Both Vav1 and Tec, however, were capable of independent engagement of a bipartite intracellular domain of the PRLR: Deletion mapping studies confined this interaction to residues 323 to 527 of the intracellular domain of the PRLR: Furthermore, Tec enhanced the GEF activity of Vav1 as evidenced by an increase in GTP-bound Rac1. These data would suggest a pivotal function for the formation of a Tec/Vav1/PRLr complex during PRL-driven signal transduction, given the role of Vav1 in the control of cell proliferation and the regulation of Rho family-mediated cytoskeletal alterations.
Mol Endocrinol 2001 May
PMID:Activation and association of the Tec tyrosine kinase with the human prolactin receptor: mapping of a Tec/Vav1-receptor binding site. 1132 62

The induction of a transformed cellular phenotype by viruses requires the modulation of signaling pathways through viral proteins. We show here that the phenotypic changes induced by the kaposin A protein of human herpesvirus 8 are mediated through its direct interaction with cytohesin-1, a guanine nucleotide exchange factor for ARF GTPases and regulator of integrin-mediated cell adhesion. Focus formation, stress fiber dissolution, and activation of the ERK-1/2 MAP kinase signal cascade were reverted by the cytohesin-1 E157K mutant, which is deficient in catalyzing guanine nucleotide exchange. Furthermore, liposome-embedded kaposin A specifically stimulates cytohesin-1 dependent GTP binding of myristoylated ARF1 in vitro. These results suggest a previously unknown involvement of ARF GTPases in the control of cellular functions by herpesviruses.
Mol Cell 2001 Apr
PMID:Signaling by human herpesvirus 8 kaposin A through direct membrane recruitment of cytohesin-1. 1133 6

Translation initiation factor 2 (eIF2) is a heterotrimeric protein that transfers methionyl-initiator tRNA(Met) to the small ribosomal subunit in a ternary complex with GTP. The eIF2 phosphorylated on serine 51 of its alpha subunit [eIF2(alphaP)] acts as competitive inhibitor of its guanine nucleotide exchange factor, eIF2B, impairing formation of the ternary complex and thereby inhibiting translation initiation. eIF2B is comprised of catalytic and regulatory subcomplexes harboring independent eIF2 binding sites; however, it was unknown whether the alpha subunit of eIF2 directly contacts any eIF2B subunits or whether this interaction is modulated by phosphorylation. We found that recombinant eIF2alpha (glutathione S-transferase [GST]-SUI2) bound to the eIF2B regulatory subcomplex in vitro, in a manner stimulated by Ser-51 phosphorylation. Genetic data suggest that this direct interaction also occurred in vivo, allowing overexpressed SUI2 to compete with eIF2(alphaP) holoprotein for binding to the eIF2B regulatory subcomplex. Mutations in SUI2 and in the eIF2B regulatory subunit GCD7 that eliminated inhibition of eIF2B by eIF2(alphaP) also impaired binding of phosphorylated GST-SUI2 to the eIF2B regulatory subunits. These findings provide strong evidence that tight binding of phosphorylated SUI2 to the eIF2B regulatory subcomplex is crucial for the inhibition of eIF2B and attendant downregulation of protein synthesis exerted by eIF2(alphaP). We propose that this regulatory interaction prevents association of the eIF2B catalytic subcomplex with the beta and gamma subunits of eIF2 in the manner required for GDP-GTP exchange.
Mol Cell Biol 2001 Aug
PMID:Tight binding of the phosphorylated alpha subunit of initiation factor 2 (eIF2alpha) to the regulatory subunits of guanine nucleotide exchange factor eIF2B is required for inhibition of translation initiation. 1143 58

Benzo[a]pyrene [B(a)P], a potent procarcinogen found in combustion products such as diesel exhaust and cigarette smoke, has been recently shown to activate the c-Jun NH(2)-terminal kinase 1 (JNK1) and induce caspase-3-mediated apoptosis in Hepa1c1c7 cells. However, the molecules of the signaling pathway that control the mitogen-activated protein kinase cascades induced by B(a)P and the interaction between those and apoptosis by B(a)P have not been well defined. We report here that B(a)P promoted Cdc42/Rac1, p21-activated kinase 1 (PAK1), and JNK1 activities in 293T and HeLa cells. Moreover, alpha-PAK-interacting exchange factor (alpha PIX) mRNA and its protein expression were upregulated by B(a)P. While overexpression of an active mutant of alpha PIX (DeltaCH) facilitated B(a)P-induced activation of Cdc42/Rac1, PAK1, and JNK1, overexpression of mutated alphaPIX (L383R, L384S), which lacks guanine nucleotide exchange factor activity, SH3 domain-deleted alphaPIX (Delta SH3), which lacks the ability to bind PAK, kinase-negative PAK1 (K299R), and kinase-negative SEK1 (K220A, K224L) inhibited B(a)P-triggered JNK1 activation. Interestingly, overexpression of alphaPIX (Delta CH) and a catalytically active mutant PAK1 (T423E) accelerated B(a)P-induced apoptosis in HeLa cells, whereas alphaPIX (Delta SH3), PAK1 (K299R), and SEK 1 (K220A, K224L) inhibited B(a)P-initiated apoptosis. Finally, a preferential caspase inhibitor, Z-Asp-CH2-DCB, strongly blocked the alphaPIX (Delta CH)-enhanced apoptosis in cells treated with B(a)P but did not block PAK1/JNK1 activation. Taken together, these results indicate that alphaPIX plays a crucial role in B(a)P-induced apoptosis through activation of the JNK1 pathway kinases.
Mol Cell Biol 2001 Oct
PMID:Involvement of alpha-PAK-interacting exchange factor in the PAK1-c-Jun NH(2)-terminal kinase 1 activation and apoptosis induced by benzo[a]pyrene. 1156 64

The alpha subunit of polypeptide chain initiation factor eIF2 can be phosphorylated by a number of related protein kinases which are activated in response to cellular stresses. Physiological conditions which result in eIF2 alpha phosphorylation include virus infection, heat shock, iron deficiency, nutrient deprivation, changes in intracellular calcium, accumulation of unfolded or denatured proteins and the induction of apoptosis. Phosphorylated eIF2 acts as a dominant inhibitor of the guanine nucleotide exchange factor eIF2B and prevents the recycling of eIF2 between successive rounds of protein synthesis. Extensive phosphorylation of eIF2 alpha and strong inhibition of eIF2B activity can result in the downregulation of the overall rate of protein synthesis; less marked changes may lead to alterations in the selective translation of alternative open reading frames in polycistronic mRNAs, as demonstrated in yeast. These mechanisms can provide a signal transduction pathway linking eukaryotic cellular stress responses to alterations in the control of gene expression at the translational level.
Prog Mol Subcell Biol 2001
PMID:Initiation factor eIF2 alpha phosphorylation in stress responses and apoptosis. 1157 61

Ras-GRF1 is a neuron-specific guanine nucleotide exchange factor for Ras proteins. Mice lacking Ras-GRF1 (-/-) are severely impaired in amygdala-dependent long-term synaptic plasticity and show higher basal synaptic activity at both amygdala and hippocampal synapses (Brambilla et al., 1997). In the present study we investigated the effects of Ras-GRF1 deletion on hippocampal neuronal excitability. Electrophysiological analysis of both primary cultured neurons and adult hippocampal slices indicated that Ras-GRF1-/- mice displayed neuronal hyperexcitability. Ras-GRF1-/- hippocampal neurons showed increased spontaneous activity and depolarized resting membrane potential, together with a higher firing rate in response to injected current. Changes in the intrinsic excitability of Ras-GRF1-/- neurons can entail these phenomena, suggesting that Ras-GRF1 deficiency might alter the balance between ionic conductances. In addition, we showed that mice lacking Ras-GRF1 displayed a higher seizure susceptibility following acute administration of convulsant drugs. Taken together, these results demonstrated a role for Ras-GRF1 in neuronal excitability.
Mol Cell Neurosci 2001 Dec
PMID:Involvement of CDC25Mm/Ras-GRF1-dependent signaling in the control of neuronal excitability. 1174 43


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