Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Grb2/Ash is composed of one SH2 and two SH3 domains and functions as an adapter linking tyrosine-kinase receptors and Ras in fibroblasts. The SH2 domain binds to tyrosine-phosphorylated proteins and the SH3 domain binds to protein containing proline-rich regions. However, the mechanisms of signal transduction through Grb2/Ash in hematopoietic cells are still unclear. By means of the binding experiments using the GST fusion protein including the full length Grb2/Ash, we have found that Shc and unidentified 130-kDa and 135-kDa proteins are associated with Grb2/Ash and that they are tyrosine-phosphorylated by treatment with granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (EPO) in a human leukemia cell line UT-7. We have purified the 130-kDa protein (pp 130) using GST-GRB2/Ash affinity column. The amino-acid sequence analysis showed that the pp130 was identical to the human c-cbl proto-oncogene product (c-Cbl). c-Cbl constitutively binds to the SH3 domain of Grb2/Ash both in vitro and in vivo but not to the SH2 domain of Grb2/Ash. Moreover, c-Cbl (pp 130) becomes tyrosine-phosphorylated rapidly and transiently depending on GM-CSF and EPO stimulation. However, we could not find the homologous regions with guanine nucleotide exchange factors or GTPase-activating proteins in the c-cbl gene. These findings strongly suggest that c-Cbl is implicated in the signal transduction of GM-CSF and EPO in hematopoietic cells, and c-Cbl and Grb2/Ash might also transduce a signal that is different from the signal leading to Ras regulation. Recently, we have shown that the proto-oncogene vav product (Vav) is also tyrosine-phosphorylated by treatment with GM-CSF and EPO and is constitutively associated with the SH3 domain of Grb2/Ash in UT-7. Another guanine nucleotide exchange factor Sos is also associated with Grb2/Ash in UT-7. It has been reported that Vav has guanine nucleotide exchange activity and activates Ras in vitro and in vivo. These data suggest that tyrosine kinases, the adapter Grb2/Ash, and the guanine nucleotide exchange factor Vav and Sos are members of a signaling pathway leading to Ras activation in hematopoietic cells.
Leukemia 1997 Apr
PMID:The signal transduction through Grb2/Ash in hematopoietic cells. 920 6

G-proteins mediate signal transfer from receptors to effector systems. In their guanosine 5'-triphosphate (GTP)-bound form, G-protein alpha-subunits activate effector systems. Termination of G-protein activation is achieved by the high-affinity GTPase [E.C. 3.6.1.-] of their alpha-subunits. Like GTP, inosine 5'-triphosphate (ITP) and xanthosine 5'-triphosphate (XTP) can support effector system activation. We studied the interactions of GTP, ITP, and XTP with the retinal G-protein, transducin (TD), and with G-proteins in HL-60 leukemia cell membranes. TD hydrolyzed nucleoside 5'-triphosphates (NTPs) in the order of efficacy GTP > ITP > XTP. NTPs eluted TD from rod outer segment disk membranes in the same order of efficacy. ITP and XTP competitively inhibited TD-catalyzed GTP hydrolysis. In HL-60 membranes, the chemoattractants N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) and leukotriene B4 (LTB4) effectively activated GTP and ITP hydrolysis by Gi-proteins. fMLP and LTB4 were at least 10-fold more potent activators of ITPase than of GTPase. Complement C5a effectively activated the GTPase of Gi-proteins but was only a weak stimulator of ITPase. The potency of C5a to activate GTP and ITP hydrolysis was similar. The fMLP-stimulated GTPase had a lower Km value than the fMLP-stimulated ITPase, whereas the opposite was true for the Vmax values. fMLP, C5a, and LTB4 did not stimulate XTP hydrolysis. Collectively, our data show that GTP, ITP, and XTP bind to G-proteins with different affinities, that G-proteins hydrolyze NTPs with different efficacies, and that chemoattractants stimulate GTP and ITP hydrolysis by Gi-proteins in a receptor-specific manner. On the basis of our results and the data in the literature, we put forward the hypothesis that GTP, ITP, and XTP act as differential signal amplifiers and signal sorters at the G-protein level.
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PMID:Functionally nonequivalent interactions of guanosine 5'-triphosphate, inosine 5'-triphosphate, and xanthosine 5'-triphosphate with the retinal G-protein, transducin, and with Gi-proteins in HL-60 leukemia cell membranes. 933 71

Gi3, a member of the Gi family of heterotrimeric GTP-binding proteins, regulates vesicle trafficking along both the constitutive and regulated pathways. In mast cells, specialized secretory cells which secrete a variety of inflammatory mediators by regulated exocytosis, activation of Gi3 provides a sufficient signal for exocytosis [Aridor, M., Rajmilevich, G., Beaven, M. A. & Sagi-Eisenberg, R. (1993) Science 262, 1569-1572]. Such activation can be achieved in patch-clamped or streptolysin-O (SLO)-permeabilized mast cells by a combination of Ca2+ and nonhydrolyzable analogs of GTP. In contrast, Ca2+-activated exocytosis in intact cells is Gi3 independent. We show here that overexpression of a GTPase-deficient mutant (G alpha i3Q204L), but not of the wild-type form of G alpha i3, in rat basophilic leukemia cells (RBL-2H3), a tumor analog of mucosal mast cells, resulted in marked potentiation of exocytosis and release of arachidonic acid in intact cells activated by a Ca2+ ionophore alone or in combination with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate. In contrast, exocytosis and arachidonic acid release stimulated by aggregation of the cell surface receptors for immunoglobulin E (IgE) were unaffected. These results strongly suggest that the intracellular receptor, responsible for the activation of Gi3, is a low-affinity Ca2+-binding protein that can only be activated during Ca2+ ionophore stimulation. Moreover, these results also suggest that the propagation of the Ca2+-activated and Gi3-mediated signaling pathway requires the blocking of Gi3 GTPase activity. Finally, our results indicate that release of arachidonic acid is at least one of the downstream effectors of Gi3.
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PMID:Stimulation of Ca2+-dependent exocytosis and arachidonic acid release in cultured mast cells (RBL-2H3) by a GTPase-deficient mutant of G alpha i3. 985 3

The import of proteins into the nucleus is dependent on cis-acting targeting sequences, nuclear localization signals (NLSs), and members of the nuclear transport receptor (importin-beta-like) superfamily. The most extensively characterized import pathway, often termed the classical pathway, is utilized by many basic-type (lysine-rich) NLSs and requires an additional component, importin alpha, to serve as a bridge between the NLS and the import receptor importin beta. More recently, it has become clear that a variety of proteins enter the nucleus via alternative import receptors and that their NLSs bind directly to those receptors. By using the digitonin-permeabilized cell system for protein import in vitro, we have defined the import pathway for the Rex protein of human T-cell leukemia virus type 1. Interestingly, the arginine-rich NLS of Rex uses importin beta for import but does so by a mechanism that is importin alpha independent. Based on the ability of the Rex NLS to inhibit the import of the lysine-rich NLS of T antigen and of both NLSs to be inhibited by the domain of importin alpha that binds importin beta (the IBB domain), we infer that the Rex NLS interacts with importin beta directly. In addition, and in keeping with other receptor-mediated nuclear import pathways, Rex import is dependent on the integrity of the Ran GTPase cycle. Based on these results, we suggest that importin beta can mediate the nuclear import of arginine-rich NLSs directly, or lysine-rich NLSs through the action of importin alpha.
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PMID:Importin beta can mediate the nuclear import of an arginine-rich nuclear localization signal in the absence of importin alpha. 989 Oct 56

GAP1(IP4BP) and GAP1(m) belong to the GAP1 family of Ras GTPase-activating proteins that are candidate InsP4 receptors. Here we show they are ubiquitously expressed in human tissues and are likely to have tissue-specific splice variants. Analysis by subcellular fractionation of RBL-2H3 rat basophilic leukemia cells confirms that endogenous GAP1(IP4BP) is primarily localised to the plasma membrane, whereas GAP1(m) appears localised to the cytoplasm (cytosol and internal membranes) but not the plasma membrane. Subcellular fractionation did not indicate a specific co-localisation between membrane-bound GAP1(m) and several Ca2+ store markers, consistent with the lack of co-localisation between GAP1(m) and SERCA1 upon co-expression in COS-7 cells. This difference suggests that GAP1(m) does not reside at a site where it could regulate the ability of InsP4 to release intracellular Ca2+. As GAP1(m) is primarily localised to the cytosol of unstimulated cells it may be spatially regulated in order to interact with Ras at the plasma membrane.
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PMID:Tissue-specific expression and endogenous subcellular distribution of the inositol 1,3,4,5-tetrakisphosphate-binding proteins GAP1(IP4BP) and GAP1(m). 1004 24

To investigate receptor-mediated Moloney murine leukemia virus (MoMuLV) entry, the green fluorescent protein (GFP)-tagged ecotropic receptor designated murine cationic amino acid transporter (MCAT-1) (MCAT-1-GFP) was constructed and expressed in 293 cells (293/MCAT-1-GFP). 293/MCAT-1-GFP cells displayed green fluorescence primarily at the cell membrane and supported wild-type levels of MoMuLV vector binding and transduction. Using immunofluorescence labeling and confocal microscopy, it was demonstrated that the surface envelope protein (SU) gp70 of MoMuLV virions began to appear inside cells 5 min after virus binding and was colocalized with MCAT-1-GFP. However, clathrin was not colocalized with MCAT-1-GFP, suggesting that MoMuLV entry, mediated by MCAT-1, does not involve clathrin. Double immunofluorescence labeling of SU and clathrin in 293 cells expressing untagged receptor (293/MCAT-1) gave the same results, i.e., SU and clathrin did not colocalize. In addition, we examined the transduction ability of MoMuLV vector on HeLa cells overexpressing the dominant-negative GTPase mutant of dynamin (K44A). HeLa cells overexpressing mutant dynamin have a severe block in endocytosis by the clathrin-coated-pit pathway. No significant titer difference was observed when MoMuLV vector was tranduced into HeLa cells overexpressing either wild-type or mutant dynamin, while the transduction ability of vesicular stomatitis virus glycoprotein pseudotyped vector into HeLa cells overexpressing mutant dynamin was decreased significantly. Taken together, these data suggest that MoMuLV entry does not occur through the clathrin-coated-pit-mediated endocytic pathway.
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PMID:Receptor-mediated Moloney murine leukemia virus entry can occur independently of the clathrin-coated-pit-mediated endocytic pathway. 1036 51

To investigate the regulation of the CCR1 chemokine receptor, a rat basophilic leukemia (RBL-2H3) cell line was modified to stably express epitope-tagged receptor. These cells responded to RANTES (regulated upon activation normal T expressed and secreted), macrophage inflammatory protein-1alpha, and monocyte chemotactic protein-2 to mediate phospholipase C activation, intracellular Ca(2+) mobilization and exocytosis. Upon activation, CCR1 underwent phosphorylation and desensitization as measured by diminished GTPase stimulation and Ca(2+) mobilization. Alanine substitution of specific serine and threonine residues (S2 and S3) or truncation of the cytoplasmic tail (DeltaCCR1) of CCR1 abolished receptor phosphorylation and desensitization of G protein activation but did not abolish desensitization of Ca(2+) mobilization. S2, S3, and DeltaCCR1 were also resistant to internalization, mediated greater phosphatidylinositol hydrolysis and sustained Ca(2+) mobilization, and were only partially desensitized by RANTES, relative to S1 and CCR1. To study CCR1 cross-regulation, RBL cells co-expressing CCR1 and receptors for interleukin-8 (CXCR1, CXCR2, or a phosphorylation-deficient mutant of CXCR2, 331T) were produced. Interleukin-8 stimulation of CXCR1 or CXCR2 cross-phosphorylated CCR1 and cross-desensitized its ability to stimulate GTPase activity and Ca(2+) mobilization. Interestingly, CCR1 cross-phosphorylated and cross-desensitized CXCR2, but not CXCR1. Ca(2+) mobilization by S3 and DeltaCCR1 were also cross-desensitized by CXCR1 and CXCR2 despite lack of receptor phosphorylation. In contrast to wild type CCR1, S3 and DeltaCCR1, which produced sustained signals, cross-phosphorylated and cross-desensitized responses to CXCR1 as well as CXCR2. Taken together, these results indicate that CCR1-mediated responses are regulated at several steps in the signaling pathway, by receptor phosphorylation at the level of receptor/G protein coupling and by an unknown mechanism at the level of phospholipase C activation. Moreover selective cross-regulation among chemokine receptors is, in part, a consequence of the strength of signaling (i.e. greater phosphatidylinositol hydrolysis and sustained Ca(2+) mobilization) which is inversely correlated with the receptor's susceptibility to phosphorylation. Since many chemokines activate multiple chemokine receptors, selective cross-regulation among such receptors may play a role in their immunomodulation.
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PMID:Regulation of the human chemokine receptor CCR1. Cross-regulation by CXCR1 and CXCR2. 1073 56

In a search for key molecules that prevent murine M1 leukemia cells from undergoing interleukin (IL)-6-induced differentiation into macrophages, we isolated an antisense complementary DNA (cDNA) that encodes full-length mouse MgcRac-GTPase-activating protein (GAP) through functional cloning. Forced expression of this antisense cDNA profoundly inhibited IL-6-induced differentiation of M1 cells into macrophage lineages. We also isolated a full-length human MgcRacGAP cDNA, which encodes an additional N-terminal polypeptide of 105 amino acid residues compared with the previously published human MgcRacGAP. In human HL-60 leukemic cells, overexpression of the full-length form of human MgcRacGAP alone induced growth suppression and macrophage differentiation associated with hypervacuolization and de novo expression of the myelomonocytic marker CD14. Analyses using a GAP-inactive mutant and 2 deletion mutants of MgcRacGAP indicated that the GAP activity was dispensable, but the myosin-like domain and the cysteine-rich domain were indispensable for growth suppression and macrophage differentiation. The present results indicated that MgcRacGAP plays key roles in controlling growth and differentiation of hematopoietic cells through mechanisms other than regulating Rac GTPase activity.
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PMID:MgcRacGAP is involved in the control of growth and differentiation of hematopoietic cells. 1097 56

The RGS (regulator of G-protein signalling) proteins are GTPase-activating proteins for activated Galpha subunits. We investigated the effects of protein kinase C (PKC) on RGS proteins in various T cell lines by treating them with PMA. mRNA levels of both RGS16 and tumour necrosis factor alpha (TNFalpha) were found to be up-regulated in CEM leukaemia cells in a PKC-dependent manner. Mezerein, a non-phorbol-ester activator of PKC, also elevated RGS16 and TNFalpha mRNA levels, while the specific PKC inhibitor Go6983 abrogated their expression. In view of the slower kinetics of PMA-induced RGS16 expression and the tight correlation between TNFalpha and RGS16 mRNA induction among the cell lines studied, we suggest that activation of PKC up-regulates RGS16 via TNFalpha. Indeed, addition of recombinant TNFalpha to CEM cells rapidly stimulated RGS16 mRNA expression independently of PKC. Furthermore, mobilization of calcium by A23187 and thapsigargin blocked the TNFalpha-mediated induction of RGS16, which was reversed by EGTA and by the immunosuppressants FK506 and cyclosporin A, suggesting that the calcineurin/NF-AT (nuclear factor of activated T cells) pathway may repress the up-regulation process. Our results demonstrate for the first time that activation of PKC induces RGS16 expression via TNFalpha in a calcium-sensitive manner, thereby implicating RGS16 in the regulation of T cell responses to inflammation.
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PMID:Specific induction of RGS16 (regulator of G-protein signalling 16) mRNA by protein kinase C in CEM leukaemia cells is mediated via tumour necrosis factor alpha in a calcium-sensitive manner. 1110 82

To define the role of regulators of G-protein signaling (RGS) in chemoattractant-mediated responses, RGS4 and the receptors for platelet-activating factor (PAFR), formylated peptides (FR), or interleukin-8 (CXCR1) were stably coexpressed in a rat basophilic leukemia (RBL-2H3) cell line. The data demonstrate that RGS4 inhibited responses by PAFR (i.e., phosphoinositide (PI) hydrolysis, Ca2+ mobilization) but not by FR or CXCR1. An N-terminal 33 amino acid deletion mutant of RGS4 (DeltaRGS4), deficient in GAP (GTPase activating protein) activity and plasma membrane localization, had no effect on either PAFR, FR, or CXCR1. RGS4, but not DeltaRGS4, also blocked phosphorylation of PAFR by platelet-activating factor (PAF) and, unexpectedly, by phorbol 12-myristate 13-acetate (PMA); it also blocked cross-phosphorylation by formylmethionylleucylphenylalanine (fMLP). A point mutant of RGS4 (N88S), deficient in GAP activity but not membrane localization, partially blocked PAFR phosphorylation but had no effect on PAFR-mediated PI hydrolysis and Ca2+ mobilization. Truncation of the cytoplasmic tail of PAFR (mPAFR) resulted in a loss of its susceptibility to inhibition by RGS4. Taken together, the data indicate that of the receptors studied, RGS4 selectively inhibited responses to PAFR, which preferentially couples to Gq. At the level of expression studied, RGS4 did not inhibit FR or CXCR1 which activates Gi to transduce cellular signals. Since the tail-deleted mutant of PAFR was not affected by RGS4, and RGS4 blocked homologous as well as heterologous phosphorylation of this receptor, it is possible that RGS4 interferes sterically with the cytoplasmic tail of PAFR. Thus, in addition to stimulating the GTPase activity of Galpha, RGS4 prevents G protein activation by PAFR and the homologous and heterologous phosphorylation of this receptor.
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PMID:RGS4 inhibits platelet-activating factor receptor phosphorylation and cellular responses. 1129 24


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