| 1. |
- Henriksson, M, et al.
(författare)
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Nuclear colocalization of c-myc protein and hsp70 in cells transfected with human wild-type and mutant c-myc genes
- 1992
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Ingår i: Experimental Cell Research. - 0014-4827. ; 203:2, s. 94-383
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Tidskriftsartikel (refereegranskat)abstract
- Using immunofluorescence and electron microscopy we have studied the localization of wild-type and mutant c-myc proteins transiently expressed in CV-1 cells. In agreement with our previous observations, wild-type c-myc protein accumulated in large amorphous globules in the nucleus. All mutant proteins tested accumulated in the nucleus as well, but gave rise to morphologically different inclusion bodies. Many small globules appeared in cells transfected with D145-262 (deletion of amino acids 145-262), while cells transfected with D371-412 or D414-433 generated structures looking like a fine network or like beads on a string. In addition, a particulate cytoplasmic staining appeared in some cells transfected with the wild-type gene and in cells transfected with mutants D145-262 or D414-433. Since the c-myc protein has been reported to stimulate expression of exogenous hsp70 protein, we also examined the intracellular distribution of hsp70 in the transfected cells. Double immunofluorescence microscopy revealed that hsp70 codistributed with the c-myc protein in distinct globules in the nucleus of many but not all myc-positive cells. However, the levels of hsp70 transcripts were not significantly raised compared to nontransfected and vector-transfected cells. Likewise, the levels of hsp70 protein did not vary significantly. These findings indicate that overexpression of c-myc stimulates translocation of preexisting hsp70 from the cytoplasm into the nucleus, rather than influencing hsp70 expression. Conceivably, this may represent one of several mechanisms whereby the cell deals with excessive amounts of c-myc protein.
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| 2. |
- Moskalewski, S., et al.
(författare)
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Influence of colchicine and vinblastine on the Golgi complex and matrix deposition in chondrocyte aggregates. An ultrastructural study
- 1975
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827. ; 95:2, s. 440-454
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Tidskriftsartikel (refereegranskat)abstract
- Fetal guinea-pig epiphyseal chondrocytes were isolated enzymatically, aggregated, and the aggregates maintained in organ culture. As revealed by light and electron microscopy, the cultures produced a typical cartilaginous matrix, but no calcification occurred. Exposure of aggregating cells, or preformed aggregates, to colchicine or vinblastihe at 10-5 M concentration led to disappearance of the microtubules, dissociation of the Golgi complex into single dictyosomes, and clustering of lysosomes. Thus, in treated cells the dictyosomes with accompanying vesicular structures were dispersed throughout the cytoplasm, whereas they were localized in a well-defined juxtanuclear region in control cells. The number and size of the cisternae forming a dictyosome were often reduced. Cells treated with vinblastine displayed macrotubules and an increased number of phagosomes. Both drugs reduced the deposition of intercellular matrix. In cells first exposed to either of the drugs for 2 or 5 days and then transferred to fresh medium for 3 or 6 days, the microtubules reappeared, the Golgi complex regained its normal appearance, and the amount of matrix increased. These findings are discussed in view of present concepts of the role of microtubules in cell secretion.
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| 3. |
- Tingström, Anders, et al.
(författare)
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Distribution and dynamics of cell surface-associated cellCAM 105 in cultured rat hepatocytes
- 1989
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 1090-2422 .- 0014-4827. ; 185:1, s. 132-142
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Tidskriftsartikel (refereegranskat)abstract
- The cellular location of cellCAM 105 was studied by indirect immunofluorescence microscopy of primary rat hepatocytes grown in monolayer culture. Staining corresponding to cellCAM 105 was seen both in cell-cell contact areas and on the upper surfaces of the cells. In the cell-cell contact areas the antigen was not accessible to the antibodies unless the cells were either permeabilized with detergent or incubated in a calcium-free medium. Removal of calcium from the medium caused the cells to separate from each other. Within a few minutes wide intercellular clefts were formed, and upon further incubation the cells became stellate-shaped and finally remained in contact with each other only via thin cellular processes. These processes were cellCAM 105-positive and at sites where they attached to the bodies of the contracted cells a granular fluorescence pattern appeared. After 24-48 h of culture, intercellular channels resembling bile canaliculi were sometimes formed in the hepatocyte monolayers. The membranes of these intercellular channels were stained for cellCAM 105. After culture for several days the hepatocytes lost their polygonal shape and gradually acquired a more fibroblast-like morphology. This morphological change was accompanied by a decrease in cellCAM 105-specific fluorescence, both in the cell-cell contact areas and on the free cell surfaces.
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| 4. |
- Westergren-Thorsson, G, et al.
(författare)
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TGF-beta enhances the production of hyaluronan in human lung but not in skin fibroblasts
- 1990
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827. ; 186:1, s. 5-192
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Tidskriftsartikel (refereegranskat)abstract
- Transforming growth factor-beta (TGF-beta) enhances the production of extracellular matrix components, such as type I and type III collagen, fibronectin, proteoglycans, in various cell types. The effect on hyaluronan synthesis in relation to proteoglycan synthesis has not been investigated. Human lung or skin fibroblast cultures were treated with TGF-beta in serum-free medium for various periods of time. 35SO4 or [3H]glucosamine was then added to the cultures in the absence of TGF-beta for up to 48 h. Hyaluronan and proteoglycans were isolated by ion-exchange chromatography and quantitated. TGF-beta induced a three- to fourfold increase in hyaluronan production by lung cells but had no effect on skin fibroblasts. In contrast, proteoglycan synthesis was enhanced in both cell types, although skin fibroblasts responded at lower concentrations of TGF-beta. Increased accumulation of hyaluronan was noted only in the cell medium, whereas proteoglycan accumulation was observed both in the medium and in the cell layer. The ED50 for TGF-beta on hyaluronan accumulation in lung cells was the same as that for proteoglycan accumulation, i.e., 40 pM. In skin fibroblasts the ED50 was considerably lower (4 pM). The induction time needed to attain full effect of TGF-beta was 6 h for both hyaluronan and proteoglycan synthesis. These results indicate that TGF-beta has tissue-specific effects on matrix production which may be of importance for control of cell proliferation in various disease states.
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| 5. |
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| 6. |
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| 7. |
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| 8. |
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| 9. |
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| 10. |
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| 11. |
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| 12. |
- Imreh, Gabriela, et al.
(författare)
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ER retention may play a role in sorting of the nuclear pore membrane protein POM121
- 2003
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Ingår i: Experimental Cell Research. - 0014-4827 .- 1090-2422. ; 284:2, s. 173-184
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Tidskriftsartikel (refereegranskat)abstract
- Integral membrane proteins of the nuclear envelope (NE) are synthesized on the rough endoplasmic reticulum (ER) and following free diffusion in the continuous ER/NE membrane system are targeted to their proper destinations due to interactions of specific domains with other components of the NE. By studying the intracellular distribution and dynamics of a deletion mutant of an integral membrane protein of the nuclear pores, POM121, which lacks the pore-targeting domain, we investigated if ER retention plays a role in sorting of integral membrane proteins to the nuclear envelope. A nascent membrane protein lacking sorting determinants is believed to diffuse laterally in the continuous ER/NE lipid bilayer and expected to follow vesicular traffic to the plasma membrane. The GFP-tagged deletion mutant, POM121(1-129)-GFP, specifically distributed within the ER membrane, but was completely absent from the Golgi compartment and the plasma membrane. Experiments using fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP) demonstrated that despite having very high mobility within the whole ER network (D = 0.41 +/- 0.11 mum(2)/s) POM121(1-129)-GFP was unable to exit the ER. It was also not detected in post-ER compartments of cells incubated at 15degreesC. Taken together, these experiments show that amino acids 1-129 of POM121 are able to retain GFP in the ER membrane and suggest that this retention occurs by a direct mechanism rather than by a retrieval mechanism. Our data suggest that ER retention might be important for sorting of POM121 to the nuclear pores.
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| 13. |
- Lennartsson, Johan, et al.
(författare)
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Identification of Tyr900 in the kinase domain of c-Kit as a Src-dependent phosphorylation site mediating interaction with c-Crk
- 2003
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Ingår i: Experimental Cell Research. - 0014-4827 .- 1090-2422. ; 288:1, s. 110-118
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Tidskriftsartikel (refereegranskat)abstract
- We have previously demonstrated that ligand-stimulation of c-Kit induces phosphorylation of Tyr568 and Tyr570 in the juxtamembrane region of the receptor, leading to recruitment, phosphorylation and activation of members of the Src family of tyrosine kinases. In this paper, we demonstrate that members of the Src family of tyrosine kinases are able to phosphorylate c-Kit selectively on one particular tyrosine residue, Tyr900, located in the second part of the tyrosine kinase domain. In order to identify potential docking partners of Tyr900, a synthetic phosphopeptide corresponding to the amino acid sequence surrounding Tyr900 was used as an affinity matrix. By use of MALDI-TOF mass spectrometry, CrkII was identified as a protein that specifically bound to Tyr900 in a phosphorylation dependent manner, possibly via the p85 subunit of PI3-kinase. Expression of a mutant receptor where Tyr900 had been replaced with a phenylalanine residue (Y900F) resulted in a receptor with reduced ability to phosphorylate CrkII. Together these data support a model where c-Src phosphorylates the receptor, thereby creating docking sites for SH2 domain containing proteins, leading to recruitment of Crk to the receptor.
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| 14. |
- Melhus, Håkan, et al.
(författare)
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Retinol-binding protein and transthyretin expressed in HeLa cells form a complex in the endoplasmic reticulum in both the absence and the presence of retinol
- 1991
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Ingår i: Experimental Cell Research. - 0014-4827 .- 1090-2422. ; 197:1, s. 119-124
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Tidskriftsartikel (refereegranskat)abstract
- To establish a suitable experimental system for studies of the interaction of retinol-binding protein (RBP) with transthyretin (TTR) we have expressed the corresponding cDNAs in HeLa cells. To investigate whether complex formation might occur already in the endoplasmic reticulum (ER), the C-terminal ER retention signal, KDEL, was attached to TTR. The tetrameric TTR-KDEL fusion protein was retained in the ER of HeLa cells. When RBP was co-expressed with TTR-KDEL, RBP was retained intracellularly. A cDNA-encoding purpurin, a protein which is 50% identical to RBP, was then expressed together with TTR-KDEL. Purpurin was not retained intracellularly and did not bind to TTR coupled to Sepharose. The effect of the vitamin A status on the secretion of TTR and RBP was examined. While TTR expressed alone was not retained intracellularly, TTR was retained in vitamin A-deficient cells when co-expressed with RBP. Addition of retinol stimulated rapid secretion of both proteins. These results demonstrate that TTR can form a complex with RBP in the ER. The data suggest that RBP and TTR are secreted as a complex.
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| 15. |
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| 16. |
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| 17. |
- Rennel, Emma, et al.
(författare)
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Regulation of endothelial cell differentiation and transformation by H-Ras
- 2003
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Ingår i: Experimental Cell Research. - 0014-4827 .- 1090-2422. ; 291:1, s. 189-200
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Tidskriftsartikel (refereegranskat)abstract
- Angiogenesis is regulated by growth factors which activate tyrosine kinase receptors leading to the activation of a number of intracellular signaling pathways. The specific function of H-Ras during FGF-2 stimulated endothelial cell differentiation, defined as invasive growth and formation of branching networks in fibrin gels, was investigated by using conditionally immortalized endothelial cell lines induced to express H-Ras mutants. Expression of inhibitory N17Ras did not impair differentiation in response to FGF-2 and TNF-alpha. The farnesyltransferase inhibitor FTI-277 inhibited farnesylation of Ras but did not inhibit differentiation of human microvascular endothelial cells or mouse brain endothelial cells. In contrast, activated V12Ras inhibited endothelial cell differentiation and cells displayed a transformed phenotype with an increased rate of proliferation and loss of contact inhibited growth. Furthermore, V12Ras expressing endothelial cells grew as solid tumors when injected subcutaneously into mice. Our data suggest that, in endothelial cells, H-Ras activity is not required for differentiation. However, this activity must be tightly regulated as aberrant activity can disturb the ability of endothelial cells to undergo differentiation.
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| 18. |
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| 19. |
- Siwicki, Jan Konrad, et al.
(författare)
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Telomere maintenance and cell cycle regulation in spontaneously immortalized T-cell lines from Nijmegen breakage syndrome patients
- 2003
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Ingår i: Experimental Cell Research. - 0014-4827 .- 1090-2422. ; 287:1, s. 178-189
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Tidskriftsartikel (refereegranskat)abstract
- Nijmegen breakage syndrome (NBS) is a rare genetic instability syndrome associated with a high incidence of lymphoid malignancies. The NBS1 protein has been implicated in telomere biology suggesting that cells from NBS patients might have deficient telomere maintenance capacity. In this study we characterized spontaneously immortalized T-cell lines derived from three NBS patients regarding growth characteristics, telomere biology, expression of cell-cycle regulators, and response to DNA damage to understand the role of NBS1 in the immortalization process. In all the NBS T-cell lines the acquisition of an immortal phenotype was associated with telomere length stabilization, high telomerase activity, and increased mRNA expression of the catalytic subunit of telomerase (hTERT), together with c-myc up-regulation. Our findings provide evidence that telomere length maintenance was intact in the T lymphocytes in the absence of a full-length NBS protein, presumably due to the presence of an alternatively transcribed NBS protein of 70 kDa. Normal protein expression patterns for pRb and p53 in all the immortal lines coincided with altered expression of some cell-cycle proteins as well as with an impaired G1/S arrest after gamma irradiation, despite a seemingly normal p53/p21 pathway. The here described, spontaneously immortalized NBS derived T-cell lines can be useful in future analysis of the biologic effects in the NBS.
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| 20. |
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| 21. |
- Tågerud, Sven, et al.
(författare)
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Receptor-mediated uptake of horseradish peroxidase in innervated and denervated skeletal muscle
- 1985
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 160:1, s. 95-105
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Tidskriftsartikel (refereegranskat)abstract
- The in vitro uptake of [3H]inulin and horseradish peroxidase (HRP) has been studied in innervated and 6 days denervated extensor digitorum longus muscle of the mouse. Both markers were taken up at a higher rate in denervated muscle. The increase in uptake after denervation was, however, larger for HRP than for [3H]inulin. After 2 h incubation at 37 °C, pH 7.3, in the presence of equimolar concentrations of HRP and [3H]inulin (approx. 2.1 μM), the uptake of HRP was approx. 8 times as great as the uptake of [3H]inulin in the same innervated muscles. In denervated muscle the HRP uptake was approx. 19 times as great as the [3H]inulin uptake in the same muscles. Various possible explanations of these differences in uptake have been considered and tested experimentally. [3H]Inulin uptake in skeletal muscle has previously been shown to obey bulk kinetics. The present investigation shows the HRP uptake to obey saturation kinetics. The HRP uptake shows dependency on divalent cations and is reduced if incubation is carried out at pH 6.4. The uptake of HRP, when used at a low, non-saturating concentration (10 μg/ml approx. 0.25 μM), is inhibited 60% by yeast mannan (0.1 mg/ml), ribonuclease B (0.1 mg/ml, approx. 7.4 μM), mannose (30 mM), monodansylcadaverine (1 mM), chloroquine (100 μM), trifluoperazine (25 μM) or maleic acid (2 mM). It is concluded that HRP is taken up in innervated and denervated skeletal muscle by a process of receptor-mediated endocytosis and that this uptake is under neurotrophic control.
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| 22. |
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| 23. |
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| 24. |
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| 25. |
- Ageberg, Malin, et al.
(författare)
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Inhibition of geranylgeranylation mediates sensitivity to CHOP-induced cell death of DLBCL cell lines.
- 2011
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 1090-2422 .- 0014-4827. ; 317, s. 1179-1191
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Tidskriftsartikel (refereegranskat)abstract
- Prenylation is a post-translational hydrophobic modification of proteins, important for their membrane localization and biological function. The use of inhibitors of prenylation has proven to be a useful tool in the activation of apoptotic pathways in tumor cell lines. Rab geranylgeranyl transferase (Rab GGT) is responsible for the prenylation of the Rab family. Overexpression of Rab GGTbeta has been identified in CHOP refractory diffuse large B cell lymphoma (DLBCL). Using a cell line- based model for CHOP resistant DLBCL, we show that treatment with simvastatin, which inhibits protein farnesylation and geranylgeranylation, sensitises DLBCL cells to cytotoxic treatment. Treatment with the farnesyl transferase inhibitor, FTI-277, or the geranylgeranyl transferase I inhibitor, GGTI-298, indicates that the reduction in cell viability was restricted to inhibition of geranylgeranylation. In addition, treatment with BMS1, a combined inhibitor of farnesyl transferase and Rab GGT, resulted in a high cytostatic effect in WSU-NHL cells, demonstrated by reduced cell viability and decreased proliferation. Co-treatment of BMS1 or GGTI-298 with CHOP showed synergistic effects with regard to markers of apoptosis. We propose that inhibition of protein geranylgeranylation together with conventional cytostatic therapy is a potential novel strategy for treating patients with CHOP refractory DLBCL.
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| 26. |
- Ahrén, Bo
(författare)
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GLP-1 for type 2 diabetes
- 2011
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 1090-2422 .- 0014-4827. ; 317:9, s. 1239-1245
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Forskningsöversikt (refereegranskat)abstract
- Glucagon-like peptide-1 (GLP-1)-based therapy of type 2 diabetes is executed either by GLP-1 receptor agonists, which stimulate the GLP-1 receptors, or by dipeptidyl peptidase-4 (DPP-4) inhibitors, which prevent the inactivation of endogenous GLP-1 thereby increasing the concentration of endogenous active GLP-1. GLP-1 activates pancreatic receptors resulting in improved glycemia through glucose-dependent stimulation of insulin secretion and inhibition of glucagon secretion. There is also a potential beta cell preservation effect, as judged from rodent studies. GLP-1 receptors are additionally expressed in extrapancreatic tissue, having potential for the treatment to reduce body weight and to potentially have beneficial cardio- and endothelioprotective effects. Clinical trials in subjects with type 2 diabetes have shown that in periods of 12 weeks or more, these treatments reduce HbA1c by approximate to 0.8-1.1% from baseline levels of 7.7-8.5%, and they are efficient both as monotherapy and in combination therapy with metformin, sulfonylureas, thiazolidinediones or insulin. Furthermore, GLP-1 receptor agonists reduce body weight, whereas DPP-4 inhibitors are body weight neutral. The treatment is safe with very low risk for adverse events, including hypoglycaemia. GLP-1 based therapy is thus a novel and now well established therapy of type 2 diabetes, with a particular value in combination with metformin in patients who are inadequately controlled by metformin alone. (C) 2011 Elsevier Inc. All rights reserved.
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| 27. |
- Aifa, Sami, 1967-, et al.
(författare)
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A basic peptide within the juxtamembrane region is required for EGF receptor dimerization
- 2005
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 302:1, s. 108-114
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Tidskriftsartikel (refereegranskat)abstract
- The epidermal growth factor receptor (EGFR) is fundamental for normal cell growth and organ development, but has also been implicated in various pathologies, notably tumors of epithelial origin. We have previously shown that the initial 13 amino acids (P13) within the intracellular juxtamembrane region (R645-R657) are involved in the interaction with calmodulin, thus indicating an important role for this region in EGFR function. Here we show that P13 is required for proper dimerization of the receptor. We expressed either the intracellular domain of EGFR (TKJM) or the intracellular domain lacking P13 (ΔTKJM) in COS-7 cells that express endogenous EGFR. Only TKJM was immunoprecipitated with an antibody directed against the extracellular part of EGFR, and only TKJM was tyrosine phosphorylated by endogenous EGFR. Using SK-N-MC cells, which do not express endogenous EGFR, that were stably transfected with either wild-type EGFR or recombinant full-length EGFR lacking P13 demonstrated that P13 is required for appropriate receptor dimerization. Furthermore, mutant EGFR lacking P13 failed to be autophosphorylated. P13 is rich in basic amino acids and in silico modeling of the EGFR in conjunction with our results suggests a novel role for the juxtamembrane domain (JM) of EGFR in mediating intracellular dimerization and thus receptor kinase activation and function. © 2004 Elsevier Inc. All rights reserved.
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| 28. |
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| 29. |
- Almkvist, Jenny, 1971, et al.
(författare)
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Newcastle disease virus neuraminidase primes neutrophils for stimulation by galectin-3 and formyl-Met-Leu-Phe
- 2004
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Ingår i: Experimental cell research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 298:1, s. 74-82
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Tidskriftsartikel (refereegranskat)abstract
- Human neutrophils are activated by the beta-galactoside-binding lectin galectin-3, provided that the cells are primed by in vivo extravasation or by in vitro preactivation with, for example, LPS. Removal of terminal sialic acid can change neutrophil functionality and responsiveness due to exposure of underlying glycoconjugate receptors or change in surface charge. Here, we investigated whether such alteration of the cell surface carbohydrate composition can alter the responsiveness of the cells to galectin-3. Neutrophils were treated with neuraminidases (NA) of different origins: Clostridium perfringens (CP), Salmonella typhimurium, Vibrio cholerae, and Newcastle disease virus (NDV). In the presence of NDV-NA, but no other NA, the otherwise non-responding neutrophils responded readily to galectin-3 by activation of the NADPH-oxidase. The galectin-3 priming effect was inhibited by the sialidase inhibitor 2,3-dehydro-2-deoxy-N-acetyl-neuraminic acid. Earlier studies have shown that priming of the neutrophil response to galectin-3 with, for example, LPS is paralleled by degranulation of intracellular vesicles and granules and upregulation of potential galectin-3 receptors. Also, NDV-NA (but not CP-NA) treatment induced degranulation, shown as an upregulation of complement receptor 3. Since not only the galectin response but also the response to the chemoattractant fMLF was primed, NDV-NA appears to induce a general priming phenomenon, possibly due to receptor upregulation by degranulation.
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| 30. |
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| 31. |
- Andersson, Ann-Sofie, et al.
(författare)
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The effects of continuous and discontinuous groove edges on cell shape and alignment.
- 2003
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Ingår i: Experimental cell research. - 0014-4827. ; 288:1, s. 177-88
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Tidskriftsartikel (refereegranskat)abstract
- Nanofabricated model surfaces and digital image analysis of cell shape were used to address the importance of a continuous sharp edge in the alignment of cells to shallow surface grooves. The grooved model surfaces had either continuous or discontinuous edges of various depths (40-400 nm) but identical surface chemistry and groove/ridge dimensions (15 microm wide). Epithelial cells were cultured on the model surfaces for 10 and 24 h. Fluorescence microscopy combined with image analysis were used to quantify cell area and alignment and to make cell shape classifications of individual cells. The degrees of alignment of cells and the percentages of elongated cell classes increased with groove depth on samples with continuous grooves. Two main differences, with regard to cell response, were observed between the continuous and discontinuous grooved surfaces. First, significantly fewer cells aligned to surface grooves with discontinuous edges than to grooves with continuous edges. Second, there were lower percentages of the elongated cell classes on discontinuous grooves than on continuous ones. We concluded that grooved surfaces with continuous edges are more potent in aligning and inducing elongated cells. The results from the present study suggest that a mechanism of alignment involving orientation along a continuous edge is likely.
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| 32. |
- Andersson, Elin, et al.
(författare)
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Ngn2 and Nurr1 act in synergy to induce midbrain dopaminergic neurons from expanded neural stem and progenitor cells.
- 2007
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 1090-2422 .- 0014-4827. ; 313:6, s. 1172-1180
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Tidskriftsartikel (refereegranskat)abstract
- Parkinson's Disease (PD) is a debilitating motor function disorder due primarily to a loss of midbrain dopaminergic neurons and a subsequent reduction in dopaminergic innervation of the striatum. Several attempts have been made to generate dopaminergic neurons from progenitor cell populations in vitro for potential use in cell replacement therapy for PD. However, expanding cells from fetal brain with retained potential for dopaminergic differentiation has proven to be difficult. In this study, we sought to generate mesencephalic dopaminergic (mesDA) neurons from an expanded population of fetal mouse ventral midbrain (VM) progenitors through the use of retroviral gene delivery. We over-expressed Ngn2 and Nurr1, two genes present in the ventral midbrain and important for normal development of mesDA neurons, in multipassaged neurosphere-expanded midbrain progenitors. We show that over-expression of Ngn2 in these progenitors results in increased neuronal differentiation but does not promote mesDA formation. We also show that over-expression of Nurr1 alone is sufficient to generate tyrosine hydroxylase (TH) expressing cells with an immature morphology, however the cells do not express any additional markers of mesDA neurons. Overexpression of Nurr1 and Ngn2 in combination generates morphologically mature TH-expressing neurons that also express additional mesencephalic markers. (c) 2006 Elsevier Inc. All rights reserved.
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| 33. |
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| 34. |
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| 35. |
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| 36. |
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| 37. |
- Aspenström, Pontus, et al.
(författare)
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Taking Rho GTPases to the next level : the cellular functions of atypical Rho GTPases
- 2007
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 313:17, s. 3673-3679
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Forskningsöversikt (refereegranskat)abstract
- The Rho GTPases are influential regulators of signalling pathways that control vital cellular processes such as cytoskeletal dynamics, gene transcription, cell cycle progression and cell transformation. A vast majority of the studies involving Rho GTPases have been focused to the famous triad, Cdc42, Rac1 and RhoA, but this protein family actually harbours 20 members. Recently, the less known Rho GTPases have received increased attention. Many of the less studied Rho GTPases have structural, as well as, functional features which makes it pertinent to classify them as atypical Rho GTPases. This review article will focus on the critical aspects of the atypical Rho GTPases, RhoH, Wrch-1, Chp and RhoBTB. These proteins are involved in a broad spectre of biological processes, such as cytoskeletal dynamics, T-cell signalling and protein ubiquitinylation. We will also discuss the roles of atypical Rho GTPases as oncogenes or tumour suppressors, as well as their potential involvement in human diseases.
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| 38. |
- Aspenström, Pontus, et al.
(författare)
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The diaphanous-related formin DAAM1 collaborates with the Rho GTPases RhoA and Cdc42, CIP4 and Src in regulating cell morphogenesis and actin dynamics
- 2006
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 312:12, s. 2180-2194
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Tidskriftsartikel (refereegranskat)abstract
- Binding partners for the Cdc42 effector CIP4 were identified by the yeast two-hybrid system, as well as by testing potential CIP4-binding proteins in coimmunoprecipitation experiments. One of the CIP4-binding proteins, DAAM1, was characterised in more detail. DAAM1 is a ubiquitously expressed member of the mammalian diaphanous-related formins, which include proteins such as mDia1 and mDia2. DAAM1 was shown to bind to the SH3 domain of CIP4 in vivo. Ectopically expressed DAAM1 localised in dotted pattern at the dorsal side of transfected cells and the protein was accumulated in the proximity to the microtubule organising centre. Moreover, ectopic expression of DAAM1 induced a marked alteration of the cell morphology, seen as rounding up of the cells, the formation of branched protrusions as well as a reduction of stress-fibres in the transfected cells. Coimmunoprecipitation experiments demonstrated that DAAM1 bound to RhoA and Cdc42 in a GTP-dependent manner. Moreover, DAAM1 was found to interact and collaborate with the non-receptor tyrosine kinase Src in the formation of branched protrusions. Taken together, our data indicate that DAAM1 communicates with Rho GTPases, CIP4 and Src in the regulation of the signalling pathways that co-ordinate the dynamics of the actin filament system.
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| 39. |
- Aspenström, Pontus
(författare)
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The mammalian verprolin homologue WIRE participates in receptor-mediated endocytosis and regulation of the actin filament system by distinct mechanisms
- 2004
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 298:2, s. 485-498
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Tidskriftsartikel (refereegranskat)abstract
- The mammalian verprolin family consists of three family members: WIP, WIRE and CR16. WIRE was recently found to bind to WASP and N-WASP and to have roles in regulating actin dynamics downstream of the platelet-derived growth factor beta-receptor. In the current study, the WASP-binding domain of WIRE was identified, with the core of the binding motif encompassing amino acid residues 408-412. A stretch of aromatic amino acid residues close to the core motif also participates in WASP binding. Amino acid substitutions in each of these motifs abrogated WASP binding, suggesting that both motifs are involved in the binding of WIRE to WASP. Interestingly, WIRE mutants unable to bind WASP were still able to induce a reorganisation of the actin filament system, indicating that WASP did not participate in the signalling pathway that link WIRE to actin dynamics. In cells ectopically expressing WIRE, the endocytosis of the platelet-derived growth factor beta-receptor was drastically reduced. However, in contrast to the effect on the actin filament system, the WIRE-induced ablation of the receptor endocytosis required an intact WASP-binding domain. Moreover, WIRE was more efficient than WIP in inhibiting the receptor endocytosis, implicating that these two mammalian verprolins have distinct roles in mammalian cells.
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| 40. |
- Aspenström, Pontus
(författare)
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The WASP-binding protein WIRE has a role in the regulation of the actin filament system downstream of the platelet-derived growth factor receptor
- 2002
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 279:1, s. 21-33
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Tidskriftsartikel (refereegranskat)abstract
- Activation of growth factor receptors, such as platelet-derived growth factor (PDGF) receptors, has a major impact on the motile behavior of vertebrate cells. The WASP family of proteins has been recognized as important regulators of actin polymerization via the activation of the Arp2/3 complex. The activity of the WASP proteins has, in turn, been shown to be governed by a number of associated proteins, including the WASP interacting protein (WIP). This report presents a novel WIP-like protein, WIRE (for WIP-related). WIRE was shown to bind to the WH1 domain of WASP and N-WASP. WIRE was localized to actin filaments in transiently transfected PAE/PDGFRbeta cells, and in cells simultaneously expressing WIRE and WASP, WIRE relocalized WASP to actin filaments, a relocalization that required direct interaction between the two proteins. In addition, WIRE was able to bind the PDGF receptor substrate Nckbeta. PDGF treatment of cells ectopically expressing WIRE resulted in formation of peripheral protrusions composed of filopodia and lamellipodia-like structures. In cells expressing both WIRE and WASP, PDGF treatment induced a translocation of WASP to the cell margin, an effect that required the presence of WIRE. Taken together, the data presented indicate that WIRE has a role in the WASP-mediated organization of the actin cytoskeleton and that WIRE is a potential link between the activated PDGF receptor and the actin polymerization machinery.
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| 41. |
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| 42. |
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| 43. |
- Axelsson, Lena, et al.
(författare)
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Clustering of β2-integrins on human neutrophils activates dual signaling pathways to Ptdlns 3-kinase
- 2000
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827. ; 256:1, s. 257-263
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Tidskriftsartikel (refereegranskat)abstract
- The β2-integrins on leukocytes can serve as a signaling unit during cell adhesion and locomotion, and to further clarify this important property we investigated the possible mechanisms of β2-integrin-induced activation of PtdIns 3-kinase. It has previously been demonstrated that clustering of β2-integrins activates p21(ras) by a tyrosine kinase-dependent pathway, and here we show that active p21(ras) interacts with its downstream target, PtdIns 3-kinase. Engagement of β2-integrins also activates the tyrosine kinases p58(c-fgr) and p59/61(hck) and causes them to associate with the p85 subunit of PtdIns 3-kinase. These findings suggest a mechanism whereby p58(c- fgr) and p59/61(hck) are directly involved in the activation of PtdIns 3- kinase. No coupling between p58(c-fgr) and p59/61(hck) could be detected; hence these kinases probably trigger independent but parallel signals to PtdIns 3-kinase. The effect of β2-integrin clustering on PtdIns 3-kinase activity was monitored as the activation of protein kinase B (PKB). Stimulation of PKB by β2-integrins was abolished by genistein and wortmannin but not by using methyl transferase inhibitors to abrogate the influence of p21(ras)-related proteins. Thus, even if PtdIns 3-kinase is not activated by p21(ras), it can maintain full enzyme activity due to the mentioned interaction with p58(c-fgr) or p59/61(hck). These tyrosine kinases apparently activate similar pathways that operate in parallel and therefore have the potential to substitute for each other in mediating adhesion and regulating cell locomotion. (C) 2000 Academic Press.
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| 44. |
- Babakov, V.N., et al.
(författare)
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RelA/NF-?B transcription factor associates with a-actinin-4
- 2008
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 314:5, s. 1030-1038
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Tidskriftsartikel (refereegranskat)abstract
- The NF-?B/RelA family of transcription factors regulates inducible transcription of a large number of genes in response to diverse stimuli. Little is known, however, about the location of NF-?B in the cytoplasm and the transport mechanism to the nucleus. We found that NF-?B is associated with the actin-binding protein a-actinin-4. NF-?B and a-actinin-4 co-localized along actin stress fibers and in membrane lamellae in A431 cells. After a 30-min stimulation with EGF or TNF-a, a-actinin-4 and p65 were found in the nucleus. Disruption of cytoskeleton by cytochalasin D prior to treatment with TNF-a led to increase of p65 nuclear translocation. Antibodies to p65 subunit of NF-?B co-immunoprecipitated a-actinin-4 from A431 cell lysates and nuclear extracts, but a-actinin-1 and ß-actin were not found in the precipitates. Affinity chromatography experiments displayed that p65 and p50 subunits of NF-?B can bind to matrix-bound chicken gizzard a-actinin. We suggest that the a-actinin-4 is important for the NF-?B nuclear translocation and its functions inside the nucleus. © 2007 Elsevier Inc. All rights reserved.
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| 45. |
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| 46. |
- Baniol, Marion, et al.
(författare)
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Identification and characterization of distinct cell cycle stages in cardiomyocytes using the FUCCI transgenic system
- 2021
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 408:2
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Tidskriftsartikel (refereegranskat)abstract
- Understanding the regulatory mechanism by which cardiomyocyte proliferation transitions to endoreplication and cell cycle arrest during the neonatal period is crucial for identifying proproliferative factors and developing regenerative therapies. We used a transgenic mouse model based on the fluorescent ubiquitination-based cell cycle indicator (FUCCI) system to isolate and characterize cycling cardiomyocytes at different cell cycle stages at a single-cell resolution. Single-cell transcriptome analysis of cycling and noncycling cardiomyocytes was performed at postnatal days 0 (P0) and 7 (P7). The FUCCI system proved to be efficient for the identification of cycling cardiomyocytes with the highest mitotic activity at birth, followed by a gradual decline in the number of cycling and mitotic cardiomyocytes during the neonatal period. Cardiomyocytes showed premature cell cycle exit at G1/S shortly after birth and delayed G1/S progression during endoreplication at P7. Single-cell RNA-seq confirmed previously described signaling pathways involved in cardiomyocyte proliferation (Erbb2 and Hippo/YAP), and maturation-related transcriptional changes during postnatal development, including the metabolic switch from glycolysis to fatty acid oxidation in cardiomyocytes. Importantly, we generated transcriptional profiles specific to cell division and endoreplication in cardiomyocytes at different developmental stages that may facilitate the identification of genes important for adult cardiomyocyte proliferation and heart regeneration. In conclusion, the FUCCI mouse provides a valuable system to study cardiomyocyte cell cycle activity at single cell resolution that can help to decipher the switch from cardiomyocyte proliferation to endoreplication, and to revert this process to facilitate endogenous repair.
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| 47. |
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| 48. |
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| 49. |
- Bauerschmidt, Christina, et al.
(författare)
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Cohesin phosphorylation and mobility of SMC1 at ionizing radiation-induced DNA double-strand breaks in human cells
- 2011
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 317:3, s. 330-337
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Tidskriftsartikel (refereegranskat)abstract
- Cohesin, a hetero-tetrameric complex of SMC1, SMC3, Rad21 and Scc3, associates with chromatin after mitosis and holds sister chromatids together following DNA replication. Following DNA damage, cohesin accumulates at and promotes the repair of DNA double-strand breaks. In addition, phosphorylation of the SMC1/3 subunits contributes to DNA damage-induced cell cycle checkpoint regulation. The aim of this study was to determine the regulation and consequences of SMC1/3 phosphorylation as part of the cohesin complex. We show here that the ATM-dependent phosphorylation of SMC1 and SMC3 is mediated by H2AX, 53BP1 and MDC1. Depletion of RAD21 abolishes these phosphorylations, indicating that only the fully assembled complex is phosphorylated. Comparison of wild type SMC1 and SMC1S966A in fluorescence recovery after photo-bleaching experiments shows that phosphorylation of SMC1 is required for an increased mobility after DNA damage in G2-phase cells, suggesting that ATM-dependent phosphorylation facilitates mobilization of the cohesin complex after DNA damage.
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| 50. |
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