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Resultados filtrados por Publicador: American Society for Biochemistry and Molecular Biology

‣ Inositol 1,4,5-Trisphosphate 3-Kinase-A Is a New Cell Motility-promoting Protein That Increases the Metastatic Potential of Tumor Cells by Two Functional Activities*

Windhorst, Sabine; Fliegert, Ralf; Blechner, Christine; Möllmann, Katharina; Hosseini, Zara; Günther, Thomas; Eiben, Maike; Chang, Lydia; Lin, Hong-Ying; Fanick, Werner; Schumacher, Udo; Brandt, Burkhard; Mayr, Georg W.
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Cellular migration is an essential prerequisite for metastatic dissemination of cancer cells. This study demonstrates that the neuron/testis-specific F-actin-targeted inositol 1,4,5-trisphosphate 3-kinase-A (ITPKA) is ectopically expressed in different human tumor cell lines and during tumor progression in the metastatic tumor model Balb-neuT. High expression of ITPKA increases invasive migration in vitro and metastasis in a xenograft SCID mouse model. Mechanistic studies show that ITPKA promotes migration of tumor cells by two different mechanisms as follows: growth factor independently high levels of ITPKA induce the formation of large cellular protrusions by directly modulating the actin cytoskeleton. The F-actin binding activity of ITPKA stabilizes and bundles actin filaments and thus increases the levels of cellular F-actin. In growth factor-stimulated cells, the catalytically active domain enhances basal ITPKA-induced migration by activating store-operated calcium entry through production of inositol 1,3,4,5-tetrakisphosphate and subsequent inhibition of inositol phosphate 5-phosphatase. These two functional activities of ITPKA stimulating tumor cell migration place the enzyme among the potential targets of anti-metastatic therapy.

‣ Dynamic Changes in the Subcellular Distribution of Gpd1p in Response to Cell Stress*

Jung, Sunhee; Marelli, Marcello; Rachubinski, Richard A.; Goodlett, David R.; Aitchison, John D.
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Gpd1p is a cytosolic NAD+-dependent glycerol 3-phosphate dehydrogenase that also localizes to peroxisomes and plays an essential role in the cellular response to osmotic stress and a role in redox balance. Here, we show that Gpd1p is directed to peroxisomes by virtue of an N-terminal type 2 peroxisomal targeting signal (PTS2) in a Pex7p-dependent manner. Significantly, localization of Gpd1p to peroxisomes is dependent on the metabolic status of cells and the phosphorylation of aminoacyl residues adjacent to the targeting signal. Exposure of cells to osmotic stress induces changes in the subcellular distribution of Gpd1p to the cytosol and nucleus. This behavior is similar to Pnc1p, which is coordinately expressed with Gpd1p, and under conditions of cell stress changes its subcellular distribution from peroxisomes to the nucleus where it mediates chromatin silencing. Although peroxisomes are necessary for the β-oxidation of fatty acids in yeast, the localization of Gpd1p to peroxisomes is not. Rather, shifts in the distribution of Gpd1p to different cellular compartments in response to changing cellular status suggests a role for Gpd1p in the spatial regulation of redox potential, a process critical to cell survival, especially under the complex stress conditions expected to occur in the wild.

‣ Protein Kinase D Controls Actin Polymerization and Cell Motility through Phosphorylation of Cortactin*

Eiseler, Tim; Hausser, Angelika; De Kimpe, Line; Van Lint, Johan; Pfizenmaier, Klaus
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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We here identify protein kinase D (PKD) as an upstream regulator of the F-actin-binding protein cortactin and the Arp actin polymerization machinery. PKD phosphorylates cortactin in vitro and in vivo at serine 298 thereby generating a 14-3-3 binding motif. In vitro, a phosphorylation-deficient cortactin-S298A protein accelerated VCA-Arp-cortactin-mediated synergistic actin polymerization and showed reduced F-actin binding, indicative of enhanced turnover of nucleation complexes. In vivo, cortactin co-localized with the nucleation promoting factor WAVE2, essential for lamellipodia extension, in the actin polymerization zone in Heregulin-treated MCF-7 cells. Using a 3-dye FRET-based approach we further demonstrate that WAVE2-Arp and cortactin prominently interact at these structures. Accordingly, cortactin-S298A significantly enhanced lamellipodia extension and directed cell migration. Our data thus unravel a previously unrecognized mechanism by which PKD controls cancer cell motility.

‣ Phospho-regulated ACAP4-Ezrin Interaction Is Essential for Histamine-stimulated Parietal Cell Secretion*

Ding, Xia; Deng, Hui; Wang, Dongmei; Zhou, Jiajia; Huang, Yuejia; Zhao, Xuannv; Yu, Xue; Wang, Ming; Wang, Fengsong; Ward, Tarsha; Aikhionbare, Felix; Yao, Xuebiao
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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The ezrin-radixin-moesin proteins provide a regulated linkage between membrane proteins and the cortical cytoskeleton and also participate in signal transduction pathways. Ezrin is localized to the apical membrane of parietal cells and couples the protein kinase A activation cascade to the regulated HCl secretion. Our recent proteomic study revealed a protein complex of ezrin-ACAP4-ARF6 essential for volatile membrane remodeling (Fang, Z., Miao, Y., Ding, X., Deng, H., Liu, S., Wang, F., Zhou, R., Watson, C., Fu, C., Hu, Q., Lillard, J. W., Jr., Powell, M., Chen, Y., Forte, J. G., and Yao, X. (2006) Mol. Cell Proteomics 5, 1437–1449). However, knowledge of whether ACAP4 physically interacts with ezrin and how their interaction is integrated into membrane-cytoskeletal remodeling has remained elusive. Here we provide the first evidence that ezrin interacts with ACAP4 in a protein kinase A-mediated phosphorylation-dependent manner through the N-terminal 400 amino acids of ACAP4. ACAP4 locates in the cytoplasmic membrane in resting parietal cells but translocates to the apical plasma membrane upon histamine stimulation. ACAP4 was precipitated with ezrin from secreting but not resting parietal cell lysates, suggesting a phospho-regulated interaction. Indeed...

‣ Phosphatidylcholine Biosynthesis during Neuronal Differentiation and Its Role in Cell Fate Determination*

Marcucci, Hebe; Paoletti, Luciana; Jackowski, Suzanne; Banchio, Claudia
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Neuronal differentiation is characterized by neuritogenesis and neurite outgrowth, processes that are dependent on membrane biosynthesis. Thus, the production of phosphatidylcholine (PtdCho), the major membrane phospholipid, should be stimulated during neuronal differentiation. We demonstrate that during retinoic acid (RA)-induced differentiation of Neuro-2a cells, PtdCho synthesis was promoted by an ordered and sequential activation of choline kinase α (CKα) and choline cytidylyltransferase α (CCTα). Early after RA stimulation, the increase in PtdCho synthesis is mainly governed by the biochemical activation of CCTα. Later, the transcription of CKα- and CCTα-encoding genes was induced. Both PtdCho biosynthesis and neuronal differentiation are dependent on ERK activation. A novel mechanism is proposed by which PtdCho biosynthesis is coordinated during neuronal differentiation. Enforced expression of either CKα or CCTα increased the rate of synthesis and the amount of PtdCho, and these cells initiated differentiation without RA stimulation, as evidenced by cell morphology and the expression of genes associated with neuritogenesis. The differentiation resulting from enforced expression of CCTα or CKα was dependent on persistent ERK activation. These results indicate that elevated PtdCho synthesis could mimic the RA signals and thus determine neuronal cell fate. Moreover...

‣ Prostaglandin E2/EP1 Signaling Pathway Enhances Intercellular Adhesion Molecule 1 (ICAM-1) Expression and Cell Motility in Oral Cancer Cells*

Yang, Shun-Fa; Chen, Mu-Kuan; Hsieh, Yih-Shou; Chung, Tsung-Te; Hsieh, Yi-Hsien; Lin, Chiao-Wen; Su, Jen-Liang; Tsai, Ming-Hsui; Tang, Chih-Hsin
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Oral squamous cell carcinoma has a striking tendency to migrate and metastasize. Cyclooxygenase (COX)-2, the inducible isoform of prostaglandin (PG) synthase, has been implicated in tumor metastasis. However, the effects of COX-2 on human oral cancer cells are largely unknown. We found that overexpression of COX-2 or exogenous PGE2 increased migration and intercellular adhesion molecule 1 (ICAM)-1 expression in human oral cancer cells. Using pharmacological inhibitors, activators, and genetic inhibition of EP receptors, we discovered that the EP1 receptor, but not other PGE receptors, is involved in PGE2-mediated cell migration and ICAM-1 expression. PGE2-mediated migration and ICAM-1 up-regulation were attenuated by inhibitors of protein kinase C (PKC)δ, and c-Src. Activation of the PKCδ, c-Src, and AP-1 signaling pathway occurred after PGE2 treatment. PGE2-induced expression of ICAM-1 and migration activity were inhibited by a specific inhibitor, siRNA, and mutants of PKCδ, c-Src, and AP-1. In addition, migration-prone sublines demonstrated that cells with increased migration ability had higher expression of COX-2 and ICAM-1. Taken together, these results indicate that the PGE2 and EP1 interaction enhanced migration of oral cancer cells through an increase in ICAM-1 production.

‣ Identification of a Novel Cell Death Receptor Mediating IGFBP-3-induced Anti-tumor Effects in Breast and Prostate Cancer*

Ingermann, Angela R.; Yang, Yong-Feng; Han, Jinfeng; Mikami, Aki; Garza, Amanda E.; Mohanraj, Lathika; Fan, Lingbo; Idowu, Michael; Ware, Joy L.; Kim, Ho-Seong; Lee, Dae-Yeol; Oh, Youngman
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Insulin-like growth factor-binding protein-3 (IGFBP-3), a major regulator of endocrine actions of IGFs, is a p53-regulated potent apoptotic factor and is significantly suppressed in a variety of cancers. Recent epidemiologic studies suggest that IGFBP-3 contributes to cancer risk protection in a variety of cancers, and a polymorphic variation of IGFBP-3 influences cancer risk, although other studies vary in their conclusions. Some antiproliferative actions of IGFBP-3 have been reported to be independent of IGFs, but the precise biochemical/molecular mechanisms of IGF-independent, antiproliferative actions of IGFBP-3 are largely unknown. Here we report a new cell death receptor, IGFBP-3R, that is a single-span membrane protein and binds specifically to IGFBP-3 but not other IGFBP species. Expression analysis of IGFBP-3 and IGFBP-3R indicates that the IGFBP-3/IGFBP-3R axis is impaired in breast and prostate cancer. We also provide evidence for anti-tumor effect of IGFBP-3R in vivo using prostate and breast cancer xenografts in athymic nude mice. Further in vitro studies demonstrate that IGFBP-3R mediates IGFBP-3-induced caspase-8-dependent apoptosis in various cancer cells. Knockdown of IGFBP-3R attenuated IGFBP-3-induced caspase activities and apoptosis...

‣ Sphingolipid-mediated Inhibition of Apoptotic Cell Clearance by Alveolar Macrophages*

Petrusca, Daniela N.; Gu, Yuan; Adamowicz, Jeremy J.; Rush, Natalia I.; Hubbard, Walter C.; Smith, Patricia A.; Berdyshev, Evgeni V.; Birukov, Konstantin G.; Lee, Chao-Hung; Tuder, Rubin M.; Twigg, Homer L.; Vandivier, R. William; Petrache, Irina
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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A decreased clearance of apoptotic cells (efferocytosis) by alveolar macrophages (AM) may contribute to inflammation in emphysema. The up-regulation of ceramides in response to cigarette smoking (CS) has been linked to AM accumulation and increased detection of apoptotic alveolar epithelial and endothelial cells in lung parenchyma. We hypothesized that ceramides inhibit the AM phagocytosis of apoptotic cells. Release of endogenous ceramides via sphingomyelinase or exogenous ceramide treatments dose-dependently impaired apoptotic Jurkat cell phagocytosis by primary rat or human AM, irrespective of the molecular species of ceramide. Similarly, in vivo augmentation of lung ceramides via intratracheal instillation in rats significantly decreased the engulfment of instilled target apoptotic thymocytes by resident AM. The mechanism of ceramide-induced efferocytosis impairment was dependent on generation of sphingosine via ceramidase. Sphingosine treatment recapitulated the effects of ceramide, dose-dependently inhibiting apoptotic cell clearance. The effect of ceramide on efferocytosis was associated with decreased membrane ruffle formation and attenuated Rac1 plasma membrane recruitment. Constitutively active Rac1 overexpression rescued AM efferocytosis against the effects of ceramide. CS exposure significantly increased AM ceramides and recapitulated the effect of ceramides on Rac1 membrane recruitment in a sphingosine-dependent manner. Importantly...

‣ Low Cholesterol Triggers Membrane Microdomain-dependent CD44 Shedding and Suppresses Tumor Cell Migration*

Murai, Toshiyuki; Maruyama, Yuusuke; Mio, Kazuhiro; Nishiyama, Hidetoshi; Suga, Mitsuo; Sato, Chikara
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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CD44 is a cell surface adhesion molecule for hyaluronan and is implicated in tumor invasion and metastasis. Proteolytic cleavage of CD44 plays a critical role in the migration of tumor cells and is regulated by factors present in the tumor microenvironment, such as hyaluronan oligosaccharides and epidermal growth factor. However, molecular mechanisms underlying the proteolytic cleavage on membranes remain poorly understood. In this study, we demonstrated that cholesterol depletion with methyl-β-cyclodextrin, which disintegrates membrane lipid rafts, enhances CD44 shedding mediated by a disintegrin and metalloproteinase 10 (ADAM10) and that cholesterol depletion disorders CD44 localization to the lipid raft. We also evaluated the effect of long term cholesterol reduction using a statin agent and demonstrated that statin enhances CD44 shedding and suppresses tumor cell migration on a hyaluronan-coated substrate. Our results indicate that membrane lipid organization regulates CD44 shedding and propose a possible molecular mechanism by which cholesterol reduction might be effective for preventing and treating the progression of malignant tumors.

‣ Cell-based Analysis of Structure-Function Activity of Threonine Aspartase 1*

Bier, Carolin; Knauer, Shirley K.; Klapthor, Alexander; Schweitzer, Andrea; Rekik, Alexander; Krämer, Oliver H.; Marschalek, Rolf; Stauber, Roland H.
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Taspase1 is a threonine protease responsible for cleaving intracellular substrates. As such, (de)regulated Taspase1 function is expected not only to be vital for ordered development but may also be relevant for disease. However, the full repertoires of Taspase1 targets as well as the exact biochemical requirements for its efficient and substrate-specific cleavage are not yet resolved. Also, no cellular assays for this protease are currently available, hampering the exploitation of the (patho)biological relevance of Taspase1. Here, we developed highly efficient cell-based translocation biosensor assays to probe Taspase1 trans-cleavage in vivo. These modular sensors harbor variations of Taspase1 cleavage sites and localize to the cytoplasm. Expression of Taspase1 but not of inactive Taspase1 mutants or of unrelated proteases triggers proteolytic cleavage and nuclear accumulation of the biosensors. Employing our assay combined with scanning mutagenesis, we identified the sequence and spatial requirements for efficient Taspase1 processing in liquid and solid tumor cell lines. Collectively, our results defined an improved Taspase1 consensus recognition sequence, Q3(F/I/L/V)2D1↓G1′X2′D3′D4′, allowing the first genome-wide bioinformatic identification of the human Taspase1 degradome. Among the 27 most likely Taspase1 targets are cytoplasmic but also nuclear proteins...

‣ Knockdown of Cyclin-dependent Kinase Inhibitors Induces Cardiomyocyte Re-entry in the Cell Cycle*

Di Stefano, Valeria; Giacca, Mauro; Capogrossi, Maurizio C.; Crescenzi, Marco; Martelli, Fabio
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Proliferation of mammalian cardiomyocytes stops rapidly after birth and injured hearts do not regenerate adequately. High cyclin-dependent kinase inhibitor (CKI) levels have been observed in cardiomyocytes, but their role in maintaining cardiomyocytes in a post-mitotic state is still unknown. In this report, it was investigated whether CKI knockdown by RNA interference induced cardiomyocyte proliferation. We found that triple transfection with p21Waf1, p27Kip1, and p57Kip2 siRNAs induced both neonatal and adult cardiomyocyte to enter S phase and increased the nuclei/cardiomyocyte ratio; furthermore, a subpopulation of cardiomyocytes progressed beyond karyokynesis, as assessed by the detection of mid-body structures and by straight cardiomyocyte counting. Intriguingly, cardiomyocyte proliferation occurred in the absence of overt DNA damage and aberrant mitotic figures. Finally, CKI knockdown and DNA synthesis reactivation correlated with a dramatic change in adult cardiomyocyte morphology that may be a prerequisite for cell division. In conclusion, CKI expression plays an active role in maintaining cardiomyocyte withdrawal from the cell cycle.

‣ Dermatopontin Interacts with Fibronectin, Promotes Fibronectin Fibril Formation, and Enhances Cell Adhesion*

Kato, Aiko; Okamoto, Osamu; Ishikawa, Kazushi; Sumiyoshi, Hideaki; Matsuo, Noritaka; Yoshioka, Hidekatsu; Nomizu, Motoyoshi; Shimada, Tatsuo; Fujiwara, Sakuhei
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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We report that dermatopontin (DP), an abundant dermal extracellular matrix protein, is found in the fibrin clot and in the wound fluid, which comprise the provisional matrix at the initial stage of wound healing. DP was also found in the serum but at a lower concentration than that in wound fluid. DP co-localized with both fibrin and fibronectin on fibrin fibers and interacted with both proteins. Both normal fibroblast and HT1080 cell adhesion to the fibrin-fibronectin matrix were dose-dependently enhanced by DP, and the adhesion was mediated by α5β1 integrin. The cytoskeleton was more organized in the cells that adhered to the fibrin-fibronectin-DP complex. When incubated with DP, fibronectin formed an insoluble complex of fibronectin fibrils as visualized by electron microscopy. The interacting sites of fibronectin with DP were the first, thirteenth, and fourteenth type III repeats (III1, III13, and III14), with III13 and III14 assumed to be the major sites. The interaction between III2–3 and III12–14 was inhibited by DP, whereas the interaction between I1–5 and III12–14 was specifically and strongly enhanced by DP. Because the interaction between III2–3 and III12–14 is involved in forming a globular conformation of fibronectin...

‣ Prostate-derived Sterile 20-like Kinases (PSKs/TAOKs) Are Activated in Mitosis and Contribute to Mitotic Cell Rounding and Spindle Positioning*

Wojtala, Rachael L.; Tavares, Ignatius A.; Morton, Penny E.; Valderrama, Ferran; Thomas, N. Shaun B.; Morris, Jonathan D. H.
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Prostate-derived sterile 20-like kinases (PSKs) 1-α, 1-β, and 2 are members of the germinal-center kinase-like sterile 20 family of kinases. Previous work has shown that PSK 1-α binds and stabilizes microtubules whereas PSK2 destabilizes microtubules. Here, we have investigated the activation and autophosphorylation of endogenous PSKs and show that their catalytic activity increases as cells accumulate in G2/M and declines as cells exit mitosis. PSKs are stimulated in synchronous HeLa cells as they progress through mitosis, and these proteins are activated catalytically during each stage of mitosis. During prophase and metaphase activated PSKs are located in the cytoplasm and at the spindle poles, and during telophase and cytokinesis stimulated PSKs are present in trans-Golgi compartments. In addition, small interfering RNA (siRNA) knockdown of PSK1-α/β or PSK2 expression inhibits mitotic cell rounding as well as spindle positioning and centralization. These results show that PSK catalytic activity increases during mitosis and suggest that these proteins can contribute functionally to mitotic cell rounding and spindle centralization during cell division.

‣ Cell-Cell Membrane Fusion Induced by p15 Fusion-associated Small Transmembrane (FAST) Protein Requires a Novel Fusion Peptide Motif Containing a Myristoylated Polyproline Type II Helix*

Top, Deniz; Read, Jolene A.; Dawe, Sandra J.; Syvitski, Raymond T.; Duncan, Roy
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Background: The p15 FAST protein mediates cell-cell fusion using a 19-residue ectodomain.

‣ Matrix Metalloproteinase 9 (MMP-9)-dependent Processing of βig-h3 Protein Regulates Cell Migration, Invasion, and Adhesion*

Kim, Yeon Hyang; Kwon, Hyung-Joo; Kim, Doo-Sik
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Background: Cell migration is involved in altering the cell and matrix interface on the cell surface.

‣ Epidermal Growth Factor (EGF)-enhanced Vascular Cell Adhesion Molecule-1 (VCAM-1) Expression Promotes Macrophage and Glioblastoma Cell Interaction and Tumor Cell Invasion*

Zheng, Yanhua; Yang, Weiwei; Aldape, Kenneth; He, Jie; Lu, Zhimin
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Background: The mechanism underlying EGF receptor (EGFR) activation-promoted glioblastoma (GBM) cell invasion remains elusive.

‣ Negative Regulation of EGFR-Vav2 Signaling Axis by Cbl Ubiquitin Ligase Controls EGF Receptor-mediated Epithelial Cell Adherens Junction Dynamics and Cell Migration*

Duan, Lei; Raja, Srikumar M.; Chen, Gengsheng; Virmani, Sumeet; Williams, Stetson H.; Clubb, Robert J.; Mukhopadhyay, Chandrani; Rainey, Mark A.; Ying, Guoguang; Dimri, Manjari; Chen, Jing; Reddi, Alagarsamy L.; Naramura, Mayumi; Band, Vimla; Band, Hamid
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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The E3 ubiquitin ligase Casitas B lymphoma protein (Cbl) controls the ubiquitin-dependent degradation of EGF receptor (EGFR), but its role in regulating downstream signaling elements with which it associates and its impact on biological outcomes of EGFR signaling are less clear. Here, we demonstrate that stimulation of EGFR on human mammary epithelial cells disrupts adherens junctions (AJs) through Vav2 and Rac1/Cdc42 activation. In EGF-stimulated cells, Cbl regulates the levels of phosphorylated Vav2 thereby attenuating Rac1/Cdc42 activity. Knockdown of Cbl and Cbl-b enhanced the EGF-induced disruption of AJs and cell motility. Overexpression of constitutively active Vav2 activated Rac1/Cdc42 and reorganized junctional actin cytoskeleton; these effects were suppressed by WT Cbl and enhanced by a ubiquitin ligase-deficient Cbl mutant. Cbl forms a complex with phospho-EGFR and phospho-Vav2 and facilitates phospho-Vav2 ubiquitinylation. Cbl can also interact with Vav2 directly in a Cbl Tyr-700-dependent manner. A ubiquitin ligase-deficient Cbl mutant enhanced the morphological transformation of mammary epithelial cells induced by constitutively active Vav2; this effect requires an intact Cbl Tyr-700. These results indicate that Cbl ubiquitin ligase plays a critical role in the maintenance of AJs and suppression of cell migration through down-regulation of EGFR-Vav2 signaling.

‣ A Germ Cell-specific Gene, Prmt5, Works in Somatic Cell Reprogramming*

Nagamatsu, Go; Kosaka, Takeo; Kawasumi, Miyuri; Kinoshita, Taisuke; Takubo, Keiyo; Akiyama, Hideo; Sudo, Tetsuo; Kobayashi, Takashi; Oya, Mototsugu; Suda, Toshio
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Germ cells possess the unique ability to acquire totipotency during development in vivo as well as give rise to pluripotent stem cells under the appropriate conditions in vitro. Recent studies in which somatic cells were experimentally converted into pluripotent stem cells revealed that genes expressed in primordial germ cells (PGCs), such as Oct3/4, Sox2, and Lin28, are involved in this reprogramming. These findings suggest that PGCs may be useful for identifying factors that successfully and efficiently reprogram somatic cells into toti- and/or pluripotent stem cells. Here, we show that Blimp-1, Prdm14, and Prmt5, each of which is crucial for PGC development, have the potential to reprogram somatic cells into pluripotent stem cells. Among them, Prmt5 exhibited remarkable reprogramming of mouse embryonic fibroblasts into which Prmt5, Klf4, and Oct3/4 were introduced. The resulting cells exhibited pluripotent gene expression, teratoma formation, and germline transmission in chimeric mice, all of which were indistinguishable from those induced with embryonic stem cells. These data indicate that some of the factors that play essential roles in germ cell development are also active in somatic cell reprogramming.

‣ Nuclear Import Mechanism for Myocardin Family Members and Their Correlation with Vascular Smooth Muscle Cell Phenotype*

Nakamura, Seiji; Hayashi, Ken'ichiro; Iwasaki, Kazuhiro; Fujioka, Tomoaki; Egusa, Hiroshi; Yatani, Hirofumi; Sobue, Kenji
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
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Myocardin (Mycd), which is essential for the differentiation of the smooth muscle cell lineage, is constitutively located in the nucleus, although its family members, myocardin-related transcription factors A and B (MRTF-A/B), mostly reside in the cytoplasm and translocate to the nucleus in response to Rho signaling. The mechanism for their nuclear import is unclear. Here we investigated the mechanism for the nuclear import of Mycd family members and demonstrated any correlation between such mechanism and the phenotype of vascular smooth muscle cells (VSMCs). In cultured VSMCs, the knockdown of importin β1 inhibited the nuclear import of Mycd and MRTF-A/B. Their NH2-terminal basic domain was identified as a binding site for importin α/β1 by in vitro analyses. However, Mycd had a higher affinity for importin α/β1 than did MRTF-A/B, even in the absence of G-actin, and Mycd affinity for importin α1/β1 was stronger than for any other importin α/β1 heterodimers. The binding of Mycd to importin α/β1 was insensitive to G-actin, whereas that of MRTF-A/B was differently inhibited by G-actin. In dedifferentiated VSMCs, the levels of importins α1 and β1 were reduced concomitant with down-regulation of Mycd, serum response factor...