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1.
In the moss Ceratodon purpureus a phytochrome gene encodes a phytochrome type (PhyCer) which has a C-terminal domain homologous to the catalytic domain of eukaryotic protein kinases (PKs). PhyCer exhibits sequence conservation to serine/ threonine as well to tyrosine kinases. Since PhyCer is expressed very weakly in moss cells, to investigate the proposed PK activity of PhyCer, we overexpressed PhyCer transiently in fibroblast cells. For this purpose we made a chimeric receptor, EC-R, which consists of the extracellular, the membrane-spanning and the juxtamembrane domains of the human epidermal growth-factor receptor (EGF-R) linked to the PK catalytic domain of PhyCer (CerKin). The expression of EC-R in transiently transfected cells was confirmed with antibodies directed against the extracellular domain of EGF-R or against CerKin. Both EGF-R and EC-R were immunoprecipitated from lysates of overexpressing cells with antibodies against the extracellular domain of EGF-R. Phosphorylation experiments were performed with the immunoprecipitates and the phosphorylation products were subjected to phosphoamino acid analysis. Phosphorylation products specifically obtained with EC-R-transfected cells exhibit phosphorylation on serine and threonine residues. In EC-R transfected cells the endogenous EGF-R showed enhanced phosphorylation of serine and threonine residues compared to EGF-R immuno-precipitated from control cells. Although CerKin is closest to the catalytic domain of a protein tyrosine kinase from Dictyostelium discoideum, EC-R does not appear to phosphorylate tyrosine residues in vitro. From our data we conclude that PhyCer carries an active PK domain capable of phosphorylating serine and threonine residues.Abbreviations CerKin
protein kinase catalytic domain of PhyCer
- EC-R
chimeric receptor consisting of the extracellular, the membrane spanning and the juxtamembrane domains of the human epidermal growth factor receptor (EGF-R) linked to the protein kinase catalytic domain of PhyCer
- EGF-R
epidermal growth factor receptor
- mAb
monoclonal antibody
- PhyCer
phytochrome gene in Ceratodon encoding a phytochrome type which has a C-terminal domain homologous to the catalytic domain of eucaryotic protein kinases
- PK
protein kinase
- PVDF
polyvinyl difluoride
- Ser
serine
- Thr
threonine
- Tyr
tyrosine
Dr. Patricia Algarra was supported by the Alexander von Humboldt Foundation, Germany. This work was supported by the Deutsche Forschungsgemeinschaft (DFG), Bonn, Germany. 相似文献
2.
Association between alphavbeta6 integrin expression, elevated p42/44 kDa MAPK, and plasminogen-dependent matrix degradation in ovarian cancer 总被引:9,自引:0,他引:9
Altered expression of alphav integrins plays a critical role in tumor growth, invasion, and metastasis. In this study, we show that normal human epithelial ovarian cell line, HOSE, and ovarian cancer cell lines, OVCA 429, OVCA 433, and OVHS-1, expressed alphav integrin and associated beta1, beta3, and beta5 subunits, but only ovarian cancer cell lines OVCA 429 and OVCA 433 expressed alphavbeta6 integrin. The expression of alphavbeta6 in OVCA 429 and OVCA 433 was far higher than alphavbeta3 and alphavbeta5 integrin and correlated with high p42/p44 mitogen activated protein kinase (MAPK) activity and high secretion of high molecular weight urokinase plasminogen activator (HMW-uPA), pro-metalloproteinase 2 and 9 (pro-MMP-9 and pro-MMP-2). In contrast to HOSE and OVHS 1, OVCA 433 and OVCA 429 exhibited approximately 2-fold more plasminogen-dependent [3H]-collagen type IV degradation. Plasminogen-dependent [3H]-collagen IV degradation was inhibited by inhibitor of uPA (amiloride) and MMP (phenanthroline) and by antibodies against uPA or MMP-9 or alphavbeta6 integrin, indicating the involvement of alphavbeta6 integrin, uPA and MMP-9 in the process. The alphavbeta6 correlated increase in HMW-uPA and pro-MMP secretion could be inhibited by tyrosine kinase inhibitor genistein or the MEK 1 inhibitor U0126, consistent with a role of active p42/44 MAPK in the elevation of uPA, MMP-9, and MMP-2 secretion. Under similar conditions, genistein and U0126 inhibited plasminogen-dependent [3H]-collagen type IV degradation. These data suggest that sustained elevation of p42/44 MAPK activity may be required for the co-expression of alphavbeta6 integrin, which in turn may regulate the malignant potential of ovarian cancer cells via proteolytic mechanisms. 相似文献
3.
Fouchier F Penel C Pierre Montero M Bremond P Champion S 《European journal of cell biology》2007,86(3):143-160
Mn(2+) was found to induce adhesion of HT29-D4 adenoma carcinoma cells to fibrinogen (Fb). This was independent of the expression of the beta3 integrin subunit and involved endogenous alphavbeta6 but not alphavbeta5 integrin. Thus, addition of Mn(2+) led to a change in integrin alphavbeta6 specificity. Furthermore, Mn(2+) was found to strongly activate the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway in the HT29-D4 cell line. As a MAPK inhibitor strongly reduced the Mn(2+)-induced cell adhesion to Fb, it is suggested that a link between MAPK activation and cell adhesion to Fb exists. Both expression and activity of matrix metalloproteinase-9 (MMP-9) were enhanced by Mn(2+) and this led to Fb processing. MMP inhibitors prevented Mn(2+)-mediated cell adhesion to Fb, leading us to suggest that Mn(2+) promoted convergent changes in integrin alphavbeta6 conformation and Fb structure through activation of ERK/MAPK and MMP-9. Finally, we found that Mn(2+) and activators of the ERK pathway cooperated in HT29-D4 cell adhesion to Fb. Such a process may be involved in bone metastasis of some cancer cells. 相似文献
4.
5.
A molecular mechanism of integrin crosstalk: alphavbeta3 suppression of calcium/calmodulin-dependent protein kinase II regulates alpha5beta1 function.
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S D Blystone S E Slater M P Williams M T Crow E J Brown 《The Journal of cell biology》1999,145(4):889-897
Many cells express more than one integrin receptor for extracellular matrix, and in vivo these receptors may be simultaneously engaged. Ligation of one integrin may influence the behavior of others on the cell, a phenomenon we have called integrin crosstalk. Ligation of the integrin alphavbeta3 inhibits both phagocytosis and migration mediated by alpha5beta1 on the same cell, and the beta3 cytoplasmic tail is necessary and sufficient for this regulation of alpha5beta1. Ligation of alpha5beta1 activates the calcium- and calmodulin-dependent protein kinase II (CamKII). This activation is required for alpha5beta1-mediated phagocytosis and migration. Simultaneous ligation of alphavbeta3 or expression of a chimeric molecule with a free beta3 cytoplasmic tail prevents alpha5beta1-mediated activation of CamKII. Expression of a constitutively active CamKII restores alpha5beta1 functions blocked by alphavbeta3-initiated integrin crosstalk. Thus, alphavbeta3 inhibition of alpha5beta1 activation of CamKII is required for its role in integrin crosstalk. Structure-function analysis of the beta3 cytoplasmic tail demonstrates a requirement for Ser752 in beta3-mediated suppression of CamKII activation, while crosstalk is independent of Tyr747 and Tyr759, implicating Ser752, but not beta3 tyrosine phosphorylation in initiation of the alphavbeta3 signal for integrin crosstalk. 相似文献
6.
Ylipalosaari M Thomas GJ Nystrom M Salhimi S Marshall JF Huotari V Tervahartiala T Sorsa T Salo T 《Experimental cell research》2005,309(2):273-283
The integrin alphavbeta6, a receptor for fibronectin, vitronectin, tenascin and TGF-beta latency-associated peptide (LAP), is not detectable on normal oral epithelium but is neo-expressed in oral squamous cell carcinomas (OSCC) and epithelial dysplasia. Previously it has been shown that alphavbeta6 integrin can up-regulate MMP-3 and -9 expression in OSCC cells. Using beta6-transfected and control OSCC cells we demonstrate that alphavbeta6 integrin down-regulates MMP-13 expression at both mRNA and protein level. Although expressing less MMP-13, beta6-transfected cells were found to have similar collagenolytic activity as control cells and invade at similar levels through type I collagen. Growth of the tumour cells in organotypic culture and confocal microscopy confirmed low levels of MMP-13 in cells with high alphavbeta6 expression. Furthermore, human squamous cell carcinomas of the tongue with high expression of alphavbeta6 showed lower MMP-13 levels than carcinomas with low levels of alphavbeta6. Our results suggest that alphavbeta6 down-regulates MMP-13 expression in OSCC cells and that MMP-13 is not essential for the degradation of type I collagen by OSCC cells. 相似文献
7.
Gramoun A Shorey S Bashutski JD Dixon SJ Sims SM Heersche JN Manolson MF 《Journal of cellular biochemistry》2007,102(2):341-352
The integrin alphavbeta3 mediates cell-matrix interactions. Vitaxin(R), a humanized monoclonal antibody that blocks human and rabbit alphavbeta3 integrins, is in clinical trials for metastatic melanoma and prostate cancer. alphavbeta3 is the predominant integrin on osteoclasts, the cells responsible for bone resorption in health and disease. Here, we report the first investigation of Vitaxin's effects on osteoclast activity. Vitaxin (100-300 ng/ml) decreased total resorption by 50%, but did not alter resorptive activity per osteoclast. Vitaxin (300 ng/ml) decreased osteoclast numbers on plastic by 35% after 48 h. Similarly, attachment after 2 h was reduced by 30% when osteoclasts were incubated with Vitaxin (300 ng/ml) for 25 min prior to plating; however, the rate of fusion of osteoclast precursors in Vitaxin-treated and control groups was equal. Using time-lapse microscopy, we evaluated the effect of Vitaxin on osteoclast morphology and found a significant reduction in osteoclast planar area only when cells were pretreated with macrophage colony stimulating factor (M-CSF). Extracellular Ca(2+) and M-CSF have opposite effects on alphavbeta3 conformation. Elevation of extracellular Ca(2+) eliminated the inhibitory effect of Vitaxin on osteoclast attachment. In contrast, the effect of Vitaxin was enhanced in cells pretreated with M-CSF. This action of M-CSF was suppressed by the phosphatidylinositol 3-kinase (PI3-kinase) inhibitor wortmannin, suggesting that M-CSF increases Vitaxin's inhibitory effect by inside-out activation of alphavbeta3. In conclusion, Vitaxin decreases resorption by impairing osteoclast attachment, without affecting osteoclast formation and multinucleation. Our data also show that Vitaxin's inhibitory effects on osteoclasts can be modulated by factors known to alter the conformation of alphavbeta3. 相似文献
8.
Infection by field strains of Foot-and-mouth disease virus (FMDV) is initiated by binding to certain species of arginine-glycine-aspartic acid (RGD)-dependent integrin including alphavbeta3 and the epithelial integrin alphavbeta6. In this report we show that the integrin alphavbeta1, when expressed as a human/hamster heterodimer on transfected CHOB2 cells, is a receptor for FMDV. Virus binding and infection mediated by alphavbeta1 was inefficient in the presence of physiological concentrations of calcium and magnesium but were significantly enhanced by reagents that activate the integrin and promote ligand binding. The ability of chimeric alpha5/alphav integrin subunits, in association with the beta1 chain, to bind FMDV and mediate infection matched the ligand binding specificity of alphavbeta1, not alpha5beta1, thus providing further evidence for the receptor role of alphavbeta1. In addition, data are presented suggesting that amino acid residues near the RGD motif may be important for differentiating between the binding specificities of alphavbeta1 and alphavbeta6. 相似文献
9.
Akakura S Singh S Spataro M Akakura R Kim JI Albert ML Birge RB 《Experimental cell research》2004,292(2):403-416
Opsonization of apoptotic cells facilitates recognition by phagocytes for the rapid clearance of potentially inflammatory cellular material. The secreted glycoprotein Milk Fat Globule Factor-E8 (MFG-E8) is a member of this family of bridging molecules and is believed to bind phosphatidylserine (PS) on the dying cell, linking it to integrin receptors on the phagocyte. Here we report the characterization of a functional signaling module involving MFG-E8, alphavbeta5 integrin, and DOCK180 for the activation of Rac1. We show that MFG-E8 and DOCK180 are both expressed in phagocytic-competent primary immature dendritic cells (DCs) and DC2.4 cells, and are potently down-regulated upon DC maturation, consistent with their role in phagocytosis and antigen capture. Coexpression of MFG-E8 with alphavbeta5 integrin potentiated integrin-mediated Rac1 activation, which was abrogated by mutagenesis in the RGD motif in MFG-E8. Moreover, expression of antisense DOCK180 abrogated MFG-E8-alphavbeta5-mediated Rac activation and impaired the phagocytosis of apoptotic cells. These data demonstrate a biochemical link between an opsonin of apoptotic cells, the alphavbeta5 integrin, and the Crk-DOCK180-Rac1 pathway, and importantly, show that MFG-E8 and DOCK180 are expressed according to the functional status of the phagocyte. 相似文献
10.
Beck V Herold H Benge A Luber B Hutzler P Tschesche H Kessler H Schmitt M Geppert HG Reuning U 《The international journal of biochemistry & cell biology》2005,37(3):590-603
We have recently described that integrin alphavbeta3 upon interaction with its major extracellular matrix ligand vitronectin induces adhesion, motility, and proliferation of human ovarian cancer cells. Due to the important function of alphavbeta3 in cancer cell biology, it has been the effort of many scientific approaches to specifically target alphavbeta3-mediated cell adhesion and tumorbiological effects arising thereof by synthetic integrin antagonists. More recently, proteins of the ADAM family have been recognized as naturally occurring integrin ligands. Among those, human ADAM15 which encompasses the integrin binding RGD motif was shown to interact with integrin alphavbeta3. Thus, we investigated in human ovarian OV-MZ-6 cancer cells, expressing both ADAM15 and alphavbeta3, whether ADAM15 might affect alphavbeta3-mediated tumorbiological effects. We stably (over)expressed ADAM15 or its extracellular domain in OV-MZ-6 cells as well as respective ADAM15 mutants containing the tripeptide SGA instead of RGD. Cells (over)expressing ADAM15-RGD exhibited a significantly reduced alphavbeta3-mediated adhesion to vitronectin. Also, a significant time-dependent decline in numbers of cells cultivated on vitronectin was noticed. This effect was found to be rather due to impaired alphavbeta3-mediated cell adhesion than decreased cell proliferation rates, since de novo DNA synthesis was not significantly altered by elevated ADAM15 expression. Moreover, a substantially decreased random cellular motility was noticed as a function of ADAM15 encompassing an intact RGD motif. In conclusion, our results point to a physiological role of ADAM15 as a natural binding partner of integrin alphavbeta3 thereby loosening tumor cell adhesion to the underlying matrix and regulating tumor cell migration and invasion. 相似文献
11.
Borza CM Pozzi A Borza DB Pedchenko V Hellmark T Hudson BG Zent R 《The Journal of biological chemistry》2006,281(30):20932-20939
Exogenous soluble human alpha3 noncollagenous (NC1) domain of collagen IV inhibits angiogenesis and tumor growth. These biological functions are attributed to the binding of alpha3NC1 to integrin alphavbeta3. However, in some tumor cells that express integrin alphavbeta3, the alpha3NC1 domain does not inhibit proliferation, suggesting that integrin alphavbeta3 expression is not sufficient to mediate the anti-tumorigenic activity of this domain. Therefore, in the present study, we searched for novel binding receptors for the soluble alpha3NC1 domain in cells lacking alphavbeta3 integrin. In these cells, soluble alpha3NC1 bound integrin alpha3beta1; however, unlike alphavbeta3, alpha3beta1 integrin did not mediate cell adhesion to immobilized alpha3NC1 domain. Interestingly, in cells lacking integrin alpha3beta1, adhesion to the alpha3NC1 domain was enhanced due to activation of integrin alphavbeta3. These findings indicate that integrin alpha3beta1 is a receptor for the alpha3NC1 domain and transdominantly inhibits integrin alphavbeta3 activation. Thus integrin alpha3beta1, in conjunction with integrin alphavbeta3, modulates cellular responses to the alpha3NC1 domain, which may be pivotal in the mechanism underpinning its anti-angiogenic and anti-tumorigenic activities. 相似文献
12.
Minami Y Ikeda W Kajita M Fujito T Amano H Tamaru Y Kuramitsu K Sakamoto Y Monden M Takai Y 《The Journal of biological chemistry》2007,282(25):18481-18496
Integrin alphavbeta3, which forms focal complexes at leading edges in moving cells, is up-regulated in cancer cells and so is implicated in their invasiveness. Necl-5, originally identified as a poliovirus receptor and also up-regulated in cancer cells, colocalizes with integrin alphavbeta3 at leading edges in moving cells and enhances growth factor-induced cell movement. Here, we show that Necl-5 interacts directly, in cis, with integrin alphavbeta3, and enhances integrin alphavbeta3 clustering and focal complex formation at leading edges in NIH3T3 cells. The extracellular region of Necl-5, but not the cytoplasmic region, is necessary for its interaction with integrin alphavbeta3; however, both regions are necessary for its action. An interaction between integrin alphavbeta3 and vitronectin and PDGF-induced activation of Rac are also necessary for integrin alphavbeta3 clustering. The interaction between Necl-5 and integrin alphavbeta3 enhances PDGF-induced Rac activation, facilitating integrin alphavbeta3 clustering presumably in a feedback amplification manner. Thus, Necl-5 has a critical role in integrin alphavbeta3 clustering and focal complex formation. 相似文献
13.
Human parechovirus 1 utilizes integrins alphavbeta3 and alphavbeta1 as receptors 总被引:1,自引:0,他引:1
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Human parechovirus 1 (HPEV1) displays an arginine-glycine-aspartic acid (RGD) motif in the VP1 capsid protein, suggesting integrins as candidate receptors for HPEV1. A panel of monoclonal antibodies (MAbs) specific for integrins alphavbeta3, alphavbeta1, and alphavbeta5, which have the ability to recognize the RGD motif, and also a MAb specific for integrin alpha2beta1, an integrin that does not recognize the RGD motif, were tested on A549 cells. Our results showed that integrin alphav-specific MAb reduced infectivity by 85%. To specify which alphav integrins the virus utilizes, we tested MAbs specific to integrins alphavbeta3 and alphavbeta1 which reduced infectivity significantly, while a MAb specific for integrin alphavbeta5, as well as the MAb specific for alpha2beta1, showed no reduction. When a combination of MAbs specific for integrins alphavbeta3 and alphavbeta1 were used, virus infectivity was almost completely inhibited; this shows that integrins alphavbeta3 and alphavbeta1 are utilized by the virus. We therefore proceeded to test whether alphav integrins' natural ligands fibronectin and vitronectin had an effect on HPEV1 infectivity. We found that vitronectin reduced significantly HPEV1 infectivity, whereas a combination of vitronectin and fibronectin abolished infection. To verify the use of integrins alphavbeta3 and alphavbeta1 as HPEV1 receptors, CHO cells transfected and expressing either integrin alphavbeta3 or integrin alphavbeta1 were used. It was shown that the virus could successfully infect these cells. However, in immunoprecipitation experiments using HPEV1 virions and allowing the virus to bind to solubilized A549 cell extract, we isolated and confirmed by Western blotting the alphavbeta3 heterodimer. In conclusion, we found that HPEV1 utilises both integrin alphavbeta3 and alphavbeta1 as receptors; however, in cells that express both integrins, HPEV1 may preferentially bind integrin alphavbeta3. 相似文献
14.
Differential alphav integrin-mediated Ras-ERK signaling during two pathways of angiogenesis 总被引:5,自引:0,他引:5
Hood JD Frausto R Kiosses WB Schwartz MA Cheresh DA 《The Journal of cell biology》2003,162(5):933-943
Antagonists of alphavbeta3 and alphavbeta5 disrupt angiogenesis in response to bFGF and VEGF, respectively. Here, we show that these alphav integrins differentially contribute to sustained Ras-extracellular signal-related kinase (Ras-ERK) signaling in blood vessels, a requirement for endothelial cell survival and angiogenesis. Inhibition of FAK or alphavbeta5 disrupted VEGF-mediated Ras and c-Raf activity on the chick chorioallantoic membrane, whereas blockade of FAK or integrin alphavbeta3 had no effect on bFGF-mediated Ras activity, but did suppress c-Raf activation. Furthermore, retroviral delivery of active Ras or c-Raf promoted ERK activity and angiogenesis, which anti-alphavbeta5 blocked upstream of Ras, whereas anti-alphavbeta3 blocked downstream of Ras, but upstream of c-Raf. The activation of c-Raf by bFGF/alphavbeta3 not only depended on FAK, but also required p21-activated kinase-dependent phosphorylation of serine 338 on c-Raf, whereas VEGF-mediated c-Raf phosphorylation/activation depended on Src, but not Pak. Thus, integrins alphavbeta3 and alphavbeta5 differentially regulate the Ras-ERK pathway, accounting for distinct vascular responses during two pathways of angiogenesis. 相似文献
15.
Kim JE Jeong HW Nam JO Lee BH Choi JY Park RW Park JY Kim IS 《The Journal of biological chemistry》2002,277(48):46159-46165
betaig-h3 is a TGF-beta-induced matrix protein known to mediate the adhesion of several cell types. In this study, we found that all four of the fas-1 domains in betaig-h3 mediate MRC-5 fibroblast adhesion and that this was specifically inhibited by a function-blocking monoclonal antibody specific for the alphavbeta5 integrin. Using deletion mutants of the fourth fas-1 domain revealed the MRC-5 cell adhesion motif (denoted the YH motif) is located in amino acids 548-614. Experiments with substitution mutants showed that tyrosine 571, histidine 572, and their flanking leucine and isoleucine amino acids, which are all highly conserved in many fas-1 domains, are essential for mediating MRC-5 cell adhesion. A synthetic 18-amino acid peptide encompassing these conserved amino acids could effectively block MRC-5 cell adhesion to betaig-h3. Using HEK293 cells stably transfected with the beta5 integrin cDNA, we confirmed that the alphavbeta5 integrin is a functional receptor for the YH motif. In conclusion, we have identified a new alphavbeta5 integrin-interacting motif that is highly conserved in the fas-1 domains of many proteins. This suggests that fas-1 domain-containing proteins may perform their biological functions by interacting with integrins. 相似文献
16.
Modulation of the urokinase-type plasminogen activator receptor by the beta6 integrin subunit 总被引:6,自引:0,他引:6
Dalvi N Thomas GJ Marshall JF Morgan M Bass R Ellis V Speight PM Whawell SA 《Biochemical and biophysical research communications》2004,317(1):92-99
Over-expression of components of the urokinase system is well documented in cancer and is thought to enable tumour cells to migrate and invade. Changes in integrin expression are also a common feature of tumours and have been linked to changes in protease activity. It has been shown that the alphavbeta6 integrin is neo-expressed in a number of epithelial carcinomas and in wound healing situations. We therefore investigated whether alphavbeta6 is able to modulate a key regulator of proteolysis, the urokinase receptor. We report that epithelial cells expressing full-length alphavbeta6 exhibit decreased urokinase receptor expression and function. Furthermore, this novel modulation requires the C-terminal 11 amino acids of the cytoplasmic tail of the beta6 integrin subunit. Cells expressing alphavbeta3, however, did not affect urokinase receptor expression. De novo expression of beta6 by melanoma cells and beta3 by epithelial cells did not influence urokinase receptor expression or function, suggesting that modulation of urokinase system is both integrin subunit and cell-specific. 相似文献
17.
Nam JO Kim JE Jeong HW Lee SJ Lee BH Choi JY Park RW Park JY Kim IS 《The Journal of biological chemistry》2003,278(28):25902-25909
betaig-h3 is an extracellular matrix protein that mediates adhesion and migration of several cell types through interaction with integrins. In the present study, we tested whether betaig-h3 mediates endothelial cell adhesion and migration, thereby regulating angiogenesis. In this study, we demonstrate that not only betaig-h3 itself but also all four fas-1 domains of betaig-h3 mediate endothelial cell adhesion and migration through interaction with the alphavbeta3 integrin. We found that the alphavbeta3 integrin-interacting motif of the four fas-1 domains of betaig-h3 is the same YH motif that we reported previously to interact with alphavbeta5 integrin. The YH peptide inhibited endothelial cell adhesion and migration in a dose-dependent manner. We demonstrate that the YH peptide has anti-angiogenic activity in vitro and in vivo using an endothelial cell tube formation assay and a Matrigel plug assay, respectively. Our results reveal that betaig-h3 bears alphavbeta3 integrin-interacting motifs that mediate endothelial cell adhesion and migration and, therefore, may regulate angiogenesis. 相似文献
18.
Recent evidence implicates a central role for PI3K signalling in mediating cell survival during the process of neuronal differentiation. Although PI3K activity is stimulated by a wide range of growth factors and cytokines in different cell lines and tissues, activation of this pathway by insulin‐like growth factor I (IGF‐I) most likely represents the main survival signal during neuronal differentiation. IGF‐I is highly expressed during development of the central nervous system, and thus is a critical factor for the development and maturation of the cerebellum. Upon ligand binding, the IGF‐I receptor phosphorylates tyrosine residues in SHC and insulin receptor substrates (IRSs) initiating two main signalling cascades, the MAP kinase and the phosphatidylinositol 3‐kinase (PI3K) pathways. Activated PI3K is composed of a catalytic subunit (p110α or β) associated with one of a large family of regulatory subunits (p85α, p85β, p55γ, p55α, and p50α). To evaluate the contributions of these various regulatory subunits to neuronal differentiation, we have used antibodies specific for each of the PI3K subunits. Using these antisera, we now demonstrate that PI3K subunits are differentially regulated in cerebellar development, and that the expression level of the p55γ regulatory subunit reaches a maximum during postnatal development, decreasing thereafter to low levels in the adult cerebellum. Furthermore, our studies reveal that the distribution of the various PI3K regulatory subunits varies during development of the cerebellum. Interestingly, p55γ is expressed in both glial and neuronal cells; moreover, in Purkinje neurones, this subunit colocalises with the IGF‐IR. © 2001 John Wiley & Sons, Inc. J Neurobiol 47: 39–50, 2001 相似文献
19.
Maubant S Leroy-Dudal J Carreiras F Deslandes E Duigou F Staedel C Gauduchon P 《Cell biology international》2007,31(2):109-118
Human ovarian surface epithelium and epithelial tumors express integrin alphavbeta5, which can interact with vitronectin. In addition, in vitro acquisition of cisplatin resistance by alphavbeta3-expressing IGROV1 cells is accompanied by cell-surface expression of integrin alphavbeta5. To further explore the role of alphavbeta5 in ovarian carcinoma cells, IGROV1 cells were stably transfected with a human beta5 integrin cDNA construct, and three beta5 transfectant clones were selected for the expression of alphavbeta5 integrin at their cell surface. Despite a delayed entry in the exponential phase of growth, beta5-transfectant cells kept a proliferation ability similar to that of parental cells, while their growth rate was hindered in the presence of an anti-alphavbeta5 blocking antibody. Only simultaneous blockade of alphavbeta3 and alphavbeta5 by specific antibodies impeded the adhesion to vitronectin of beta5 transfectants and of the beta5-expressing cisplatin-resistant variant IGROV1-R10, suggesting that the two heterodimers cooperated in the regulation of this process. Cell surface expression of alphavbeta5 resulted in an attenuation of alphavbeta3-mediated migration on vitronectin. Alphavbeta5 participated to migration events in the absence of exogenous growth factors only in one transfectant clone and in IGROV1-R10 cells. Finally, the response to cisplatin was not significantly modified in beta5 transfectants when compared to IGROV1 parental cells. 相似文献