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Overexpression Studies (overexpression + studies)
Selected AbstractsSox genes regulate type 2 collagen expression in avian neural crest cellsDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 8 2006Takashi Suzuki Neural crest cells give rise to a wide variety of cell types, including cartilage cells in the cranium and neurons and glial cells in the peripheral nervous system. To examine the relationship of cartilage differentiation and neural crest differentiation, we examined the expression of Col2a1, which encodes type 2 collagen often used as a cartilage marker, and compared it with the expression of Sox transcription factor genes, which are involved in neural crest development and chondrogenesis. We found that Col2a1 is expressed in many neural crest-derived cell types along with combinations of Sox9, Sox10 and LSox5. Overexpression studies reveal the activation of Col2a1 expression by Sox9 and Sox10, and cross-regulation of these Sox genes. Luciferase assay indicates a direct activation of the Col2a1 enhancer/promoter both by Sox9 and Sox10, and this activation is further enhanced by cAMP-dependent kinase (PKA) signaling. Our study suggests that the regulatory mechanisms are similar in cartilage and neural crest differentiation. [source] Differential sensitivity in the survival of oligodendrocyte cell lines to overexpression of myelin proteolipid protein gene productsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2001Ernesto R. Bongarzone Abstract The proteolipid (PLP) gene encodes at least four proteins, including the classic PLP and DM20, which are important components of the myelin sheath, and the recently identified soma-restricted (sr) isoforms, srPLP and srDM20. The classic PLP and DM20 gene products have been implicated in oligodendrocyte survival by overexpression studies in vitro and in vivo. The classic and sr proteolipids are targeted to different cellular compartments in the oligodendrocyte, suggesting different cellular functions. Accordingly, we examined the effects of in vitro overexpression of the sr-PLP/DM20 isoforms on the survival of stably transfected, conditionally immortalized, oligodendroglial cell lines and compared this to overexpression of the classic and the jimpy-mutated proteolipids. The results indicate that overexpression of either normal or jimpy classic PLP/DM20 resulted in a dramatic reduction in the survival of the oligodendrocyte cell lines at the nonpermissive temperature, but not the COS-7 cell line, a cell line expressing the same oncogene constitutively. Survival of the oligodendrocyte cell lines was significantly less affected when either the sr-PLP/DM20 or the dopamine D-2 receptor, another cell membrane protein, was overexpressed in the cell lines. These results suggest that overexpression of the "classic" PLP or DM20 can compromise the survival of oligodendrocytes whether or not they are mutated. Furthermore, they suggest that the internal mechanisms for normal targeting of the PLP/DM20 isoforms of either the "classic" or the "sr" types influence the oligodendrocyte's ability to survive when these proteolipids are overexpressed. J. Neurosci. Res. 65:485,492, 2001. © 2001 Wiley-Liss, Inc. [source] The regulation of integrin-linked kinase in human platelets: evidence for involvement in the regulation of integrin ,2,1JOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 8 2004J. M. Stevens Summary.,Background: Activation of the platelet integrin ,2,1 is closely regulated due to the high thrombogenicity of its ligand. As a ,1 interacting kinase, ILK represents a candidate intracellular regulator of ,2,1 in human platelets. Objectives We investigated the regulation of ILK in human platelets and the role of ILK in regulating ,2,1 activation in HEL cells, a megakaryocytic cell line. Methods: An in-vitro kinase assay was used to determine the effect of platelet agonists on ILK kinase activity together with the contribution of PI3K and PKC on ILK activation. Interaction of ILK with ,1 -integrin subunits was investigated by coimmunoprecipitation and the role of ILK in regulating ,2,1 function assessed by overexpression studies in HEL cells. Results: We report that collagen and thrombin modulate ILK kinase activity in human platelets in an aggregation-independent manner. Furthermore, ILK activity is dually regulated by PI3K and PKC in thrombin-stimulated platelets and regulated by PI3K in collagen-stimulated cells. ILK associates with the ,1 -integrin subunits immunoprecipitated from platelet cell lysates, an association which increased upon collagen stimulation. Overexpression of ILK in HEL cells enhanced ,2,1 -mediated adhesion whereas overexpression of kinase-dead ILK reduced adhesion, indicating a role for this kinase in the positive regulation of ,2,1. Conclusions: Our findings that ILK regulates ,2,1 in HEL cells, is activated in platelets and associates with ,1 -integrins, raise the possibility that it may play a key role in adhesion events upon agonist stimulation of platelets. [source] Phenotypic changes associated with DYNACTIN-2 (DCTN2) over expression characterise SJSA-1 osteosarcoma cellsMOLECULAR CARCINOGENESIS, Issue 3 2006Kieran L. Bransfield Abstract DYNACTIN-2 (DCTN2) localises to chromosome 12q13-q15, a region prone to stable amplification in several cancers. Transient DCTN2 overexpression has a significant impact on cellular phenotype primarily due to disruption of the DYNEIN-dynactin motor. Changes reported include alterations of microtubule-directed movement of molecular (e.g. TP53) and organelle (e.g. Golgi) cargoes towards the nucleus, centrosome biology, cellular movement and mitosis with a potential predisposition to mitotic block and polyploidy. These changes would be expected to be of relevance to carcinogenesis. To investigate this, we report the first study of DCTN2 genomic amplification and sustained DCTN2 overexpression in cancer cells. QFMPCR was employed to characterise the extent of chromosome 12q13-q15 amplicons in SJSA-1, SJRH30, U373MG and CCF-STTG1 cancer cells. DCTN2 amplification was present in SJSA-1, U373MG and SJRH30 cells, yet was incomplete at the 5,-end in SJRH30 cells. Only SJSA-1 cells were characterised by DCTN2 overexpression on Western blot analyses. Microscopy studies distinguished SJSA-1 cells by greater DCTN2 immunofluorescence and diminished centrosome and 58K protein Golgi-marker focus compared to SJRH30 cells. Indirect evidence derived from the published work of others indicated that TP53 transport into the nucleus was unimpaired. Furthermore, we observed that SJSA-1 cells were easy to propagate. In conclusion, persistent DCTN2 overexpression can be tolerated in SJSA-1 cancer cells despite phenotypic abnormalities predicted from transient overexpression studies. This preliminary study does not support a major role for DCTN2 overexpression in carcinogenesis, although further studies would be necessary to confirm this. © 2005 Wiley-Liss, Inc. [source] | |||||||||||||