| 1. |
- Jia, Min, et al.
(författare)
-
Proteome profiling of immortalization-to-senescence transition of human breast epithelial cells identified MAP2K3 as a senescence-promoting protein which is downregulated in human breast cancer
- 2010
-
Ingår i: PROTEOMICS - Clinical Applications. - : Wiley. - 1862-8346 .- 1862-8354. ; 4:10-11, s. 816-828
-
Tidskriftsartikel (refereegranskat)abstract
- Purpose: Immortalization is one of the first changes in cells undergoing carcinogenic transformation. Proteome profiling of the immortalization-senescence transition is expected to provide insights into the molecular mechanisms of early tumorigenesis. Experimental design: 2-DE and MALDI-MS were used to identify proteins in primary human breast epithelial cells, relevant to the immortalization-senescence transition. Cell and molecular biology and immunohistochemistry were used to validate involvement of mitogen-activated protein kinase kinase 3 (MAP2K3) in the immortalization-senescence transition. Results: We identified 71 proteins whose expression changed upon induction of senescence. The identified proteins include regulators of cell growth, death, cell assembly and organization. Analysis of the network formed by the identified proteins suggested that the immortalization-to-senescence transition could affect regulators of the cell cycle, protein synthesis, transport, post-translational modifications, DNA recombination and repair, and lipid and amino acid metabolism. We observed that MAP2K3 was downregulated in immortal human breast epithelial cells and that upregulation of MAP2K3 expression promoted cell senescence. Decreased expression of MAP2K3 was observed in human breast infiltrating ductal carcinomas, as compared to non-cancerous human breast tissues. Conclusion and clinical relevance: We described a proteome profile of the immortalization-to-senescence transition for human breast epithelial cells, and identified MAP2K3 as a protein that promotes senescence in these cells.
|
|
| 2. |
- Kwiecińska, Anna, et al.
(författare)
-
Proteomic Profiling of Diffuse Large B-Cell Lymphomas
- 2018
-
Ingår i: Pathobiology. - : S. Karger AG. - 1015-2008 .- 1423-0291. ; 85:4, s. 211-219
-
Tidskriftsartikel (refereegranskat)abstract
- Objective: The aim of this study was to identify differences in proteome profiles of diffuse large B-cell lymphoma (DLBCL) of nongerminal center (non-GC) versus GC type in the search for new markers and drug targets. Methods: Six DLBCL, with 3 repeats for each, were used for the initial study by proteomics: 3 non-GC and 3 GC DLBCL cases. For immunohistochemistry, tissue microarrays were made from 31 DLBCL samples: 16 non-GC de novo lymphomas and 15 GC cases (11 transformed from follicular lymphomas and 4 de novo GC lymphomas). Proteome profiling was performed by two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. Results: Ninety-one proteins were found differentially expressed in non-GC compared to GC type. The Cytoscape tool was used for systemic analysis of proteomics data, revealing 19 subnetworks representing functions affected in non-GC versus GC types of DLBCL. Conclusion: A validation study of 3 selected proteins (BiP/Grp78, Hsp90, and cyclin B2) showed the enhanced expression in non-GC DLBCL, supporting the proteomics data.
|
|
| 3. |
- Sasaki, Takaaki, et al.
(författare)
-
Elevated Intraocular Pressure, Optic Nerve Atrophy, and Impaired Retinal Development in ODAG Transgenic Mice
- 2009
-
Ingår i: Investigative Ophthalmology and Visual Science. - : Association for Research in Vision and Ophthalmology (ARVO). - 0146-0404 .- 1552-5783. ; 50:1, s. 242-248
-
Tidskriftsartikel (refereegranskat)abstract
- PURPOSE. In an earlier study, a cDNA was cloned that showed abundant expression in the eye at postnatal day (P) 2 but was downregulated at P10; it was named ODAG (ocular development-associated gene). Its biological function was examined by generating and analyzing transgenic mice overexpressing ODAG (ODAG Tg) in the eye and by identifying ODAG-binding proteins. METHODS. Transgenic mice were generated by using the mouse Crx promoter. EGFP was designed to be coexpressed with transgenic ODAG, to identify transgene-expressing cells. Overexpression of ODAG was confirmed by Northern and Western blot analysis. IOP was measured with a microneedle technique. The eyes were macroscopically examined and histologically analyzed. EGFP expression was detected by confocal microscope. Proteins associated with ODAG were isolated by pull-down assay in conjugation with mass spectrometry. RESULTS. Macroscopically, ODAG Tg exhibited gradual protrusion of the eyeballs. The mean IOP of ODAG Tg was significantly higher than that of wild-type (WT) littermates. Histologic analysis exhibited optic nerve atrophy and impaired retinal development in the ODAG Tg eye. EGFP was expressed highly in the presumptive outer nuclear layer and weakly in the presumptive inner nuclear layer in the ODAG Tg retina. Rab6-GTPase-activating protein (Rab6-GAP) and its substrate, Rab6, were identified as ODAG-binding proteins. CONCLUSIONS. Deregulated expression of ODAG in the eye induces elevated intraocular pressure and optic nerve atrophy and impairs retinal development, possibly by interfering with the Rab6/Rab6-GAP-mediated signaling pathway. These results provide new insights into the mechanisms regulating ocular development, and ODAG Tg would be a novel animal model for human diseases caused by ocular hypertension.
|
|
| 4. |
- Souchelnytskyi, Serhiy, et al.
(författare)
-
COVID-19 engages clinical markers for the management of cancer and cancer-relevant regulators of cell proliferation, death, migration, and immune response
- 2021
-
Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11:1
-
Tidskriftsartikel (refereegranskat)abstract
- Clinical reports show that the management of cancer patients infected with SARS-CoV-2 requires modifications. Understanding of cancer-relevant mechanisms engaged by the virus is essential for the evidence-based management of cancer. The network of SARS-CoV-2 regulatory mechanisms was used to study potential engagement of oncogenes, tumor suppressors, other regulators of tumorigenesis and clinical markers used in the management of cancer patients. Our network analysis confirms links between COVID-19 and tumorigenesis that were predicted in epidemiological reports. The COVID-19 network shows the involvement of tumorigenesis regulators and clinical markers. Regulators of cell proliferation, death, migration, and the immune system were retrieved. Examples are pathways initiated by EGF, VEGF, TGF beta and FGF. The SARS-CoV-2 network engages markers for diagnosis, prognosis and selection of treatment. Intersection with cancer diagnostic signatures supports a potential impact of the virus on tumorigenesis. Clinical observations show the diversity of symptoms correlating with biological processes and types of cells engaged by the virus, e.g. epithelial, endothelial, smooth muscle, glial and immune system cells. Our results describe an extensive engagement of cancer-relevant mechanisms and clinical markers by COVID-19. Engagement by the virus of clinical markers provides a rationale for clinical decisions based on these markers.
|
|
| 5. |
- Bhaskaran, Nimesh, et al.
(författare)
-
Comparative proteome profiling of MCF10A and 184A1 human breast epithelial cells emphasized involvement of CDK4 and cyclin D3 in cell proliferation
- 2009
-
Ingår i: Proteomics Clinical Applications. - : Wiley. - 1862-8354 .- 1862-8346. ; 3:1, s. 68-77
-
Tidskriftsartikel (refereegranskat)abstract
- Acquiring high proliferation rate is crucial for carcinogenic transformation of cells. We reporthere proteome profiling of human breast epithelial cells with low (184A1) and high (MCF10A)proliferation rates.We identified 183 proteins in 184A1 and 318 proteins in MCF10A cells. Thesedatasets provide the most comprehensive proteome annotations of 184A1 and MCF10A cells.Proteins were taken for identification from 2-D gels in a systematic and unbiased way. Functionalclustering of the identified proteins showed similarities in distribution of proteins to the samefunctional domains, indicating similarities in proteomes of 184A1 and MCF10A cells. Amongobserved differences in protein expression, we validated correlation of expression of endogenouscyclin-dependent kinase 4 (CDK4), cyclin D3, cdc25B, and p38g with cell proliferation. Furthermore,down-regulation of CDK4 and cyclin D3 with specific siRNA inhibited cell proliferation,which emphasized the role of CDK4 and cyclin D3 in enhancement of cell proliferation rate ofhuman breast epithelial cells.
|
|
| 6. |
- Bhaskaran, Nimesh, et al.
(författare)
-
Novel post-translational modifications of Smad2 identified by mass spectrometry
- 2008
-
Ingår i: Central European Journal of Biology. - : Walter de Gruyter GmbH. - 1895-104X .- 1644-3632. ; 3:4, s. 359-370
-
Tidskriftsartikel (refereegranskat)abstract
- Smad2 is a crucial component of transforming growth factor-b (TGFb) signaling, and is involved in the regulation of cell proliferation,death and differentiation. Phosphorylation, ubiquitylation and acetylation of Smad2 have been found to regulate its activity. We usedmass spectrometry to search for novel post-translational modifications (PTMs) of Smad2. Peptide mass fingerprinting (PMF) indicatedthat Smad2 can be acetylated, methylated, citrullinated, phosphorylated and palmitoylated. Sequencing of selected peptides validatedmethylation at Gly122 and hydroxylation at Trp18 of Smad2. We also observed a novel, so far unidentified modification at Tyr128 andTyr151. Our observations open for further exploration of biological importance of the detected PTMs.
|
|
| 7. |
- Conrotto, Paolo, et al.
(författare)
-
Interactome of transforming growth factor-beta type I receptor (T beta RI) : Inhibition of TGF beta signaling by Epac1
- 2007
-
Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 6:1, s. 287-297
-
Tidskriftsartikel (refereegranskat)abstract
- Transforming growth factor-beta (TGF beta) is a potent regulator of cell growth, differentiation, and apoptosis. Type I TGF beta receptor (T beta RI) is the key receptor for initiation of intracellular signaling by TGF beta. Here we report proteomics-based identification of proteins that form a complex with T beta RI. Using 2D-GE and MALDI TOF mass spectrometry, we identified 16 proteins that specifically interacted with a GST-fused T beta RI Thr204Asp construct with constitutively active serine/threonine kinase. We confirmed interactions of the receptor with cAMP regulated guanine nucleotide exchange factor 1 (Epac1), alpha-spectrin, PIASy, and alpha-catenin proteins using immunoblotting. Interaction of the receptor with Epac1 required intact kinase activity of T beta RI but was not affected by deletion of cAMP-binding domain of Epac1. TGF beta 1-induced C-terminal phosphorylation of Smad2 was inhibited in vivo and in vitro in the presence of Epac1. Epac1 inhibited also TGF beta 1/T beta RI-dependent transcriptional activation, as evaluated by luciferase reporter assays. We observed that expression of Epac1 counteracted TGF beta/T beta RI-dependent decrease of cell adhesion and TGF beta/T beta RI-induced stimulation of cell migration. Thus, we have reported novel T beta RI-interacting proteins and have shown that Epac1 inhibited TGF beta-dependent regulation of cell migration and adhesion.
|
|
| 8. |
- Cunha, Sara I., et al.
(författare)
-
Exposure to EGF and 17 beta-estradiol irreversibly affects the proliferation and transformation of MCF7 cells but is not sufficient to promote tumor growth in a xenograft mouse model upon withdrawal of exposure
- 2018
-
Ingår i: International Journal of Molecular Medicine. - : SPANDIDOS PUBL LTD. - 1107-3756 .- 1791-244X. ; 42:3, s. 1615-1624
-
Tidskriftsartikel (refereegranskat)abstract
- Epidermal growth factor (EGF) and estrogen are potent regulators of breast tumorigenesis. Their short-term actions on human breast epithelial cells have been investigated extensively. However, the consequence of a long-term exposure to EGF and estrogen remains to be fully elucidated. The present study examined the effects of long-term exposure to EGF and 17 beta-estradiol on the proliferation, transformation, expression of markers of stemness, and tumorigenesis of MCF7 human breast adenocarcinoma cells. Exposure to EGF and/or 17 beta-estradiol irreversibly enhanced the proliferation rate of MCF7 cells, even following withdrawal. However, in a mouse xenograft experiment, no significant difference in tumor volume was observed between tumors derived from cells exposed to EGF, 17 beta-estradiol or EGF + 17 beta-estradiol. Immunohistochemistry performed on tumors derived from 17 beta-estradiol-exposed cells revealed reduced cell proliferation and vessel scores, according to the results obtained using Ki67 and von Willebrand factor staining, respectively. The EGF-and/or 17 beta-estradiol-treated cells exhibited an increased ratio of cluster of differentiation (CD) 44(+)/CD24(-) cells and enhanced ability to form mammospheres. Furthermore, the long-term exposure of MCF7 cells to EGF and 17 beta-estradiol altered their responsiveness to short-term stimulatory or inhibitory treatments with EGF, 17 beta-estradiol, transforming growth factor-beta 1 (TGF beta 1), Iressa and SB431542. Therefore, the findings indicated that sustained exposure of MCF7 cells to EGF and/or 17 beta-estradiol resulted in enhanced cell proliferation and mammosphere formation, an increased ratio of CD 44(+)/CD24 cells, and altered responses to short-term treatments with EGF, 17 beta-estradiol, TGF beta 1, and drugs inhibiting these signaling pathways. However, this sustained exposure was not sufficient to affect tumor take or volume in a xenograft mouse model.
|
|
| 9. |
- Dubrovska, Anna, et al.
(författare)
-
Efficient enrichment of intact phosphorylated proteins by modified immobilized metal-affinity chromatography
- 2005
-
Ingår i: Proteomics. - : Wiley. - 1615-9853 .- 1615-9861. ; 5:18, s. 4678-4683
-
Tidskriftsartikel (refereegranskat)abstract
- Phosphoproteome studies are hampered by the lack of methods which allow a comprehensive and fast analysis of intact phosphoproteins. Here we describe an immobilized metal-affinity chromatography (IMAC)-based technique for the enrichment of phosphorylated proteins, which allows recovery of up to 90% of phosphoproteins. This technique is compatible with 2-DE and can be applied to cultured cells and tissues.
|
|
| 10. |
- Dubrovska, Anna, et al.
(författare)
-
TGFbeta1/Smad3 counteracts BRCA1-dependent repair of DNA damage
- 2005
-
Ingår i: Oncogene. - : Springer Science and Business Media LLC. - 0950-9232 .- 1476-5594. ; 24:14, s. 2289-2297
-
Tidskriftsartikel (refereegranskat)abstract
- Inactivation of the BRCA1 gene has been found to confer susceptibility to early-onset familial breast and ovarian cancers. BRCA1 regulates DNA repair, chromatin remodeling and affects gene transcription. Transforming growth factor-beta (TGFbeta) is a potent regulator of growth, apoptosis and invasiveness of tumor cells, including breast cancer cells. Here we show that Smad3 which is a component of the TGFbeta signaling pathway, forms a complex with BRCA1 in vitro and in vivo. The interaction is mediated by the MH1 domain of Smad3 and the C-terminal part of BRCA1. We observed a co-localization of Smad3 and BRCA1 in nuclear complexes. We also found that TGFbeta1/Smad3 counteracted BRCA1-dependent repair of DNA double-strand breaks in human breast epithelial cells, as evaluated by BRCA1 nuclear foci formation, single-cell gel electrophoresis and cell survival assays. Thus, TGFbeta1/Smad3 suppresses BRCA1-dependent DNA repair in response to a DNA damaging agent.
|
|
