| 1. |
- Gupta, Rajesh Kumar, et al.
(författare)
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beta 1 Integrins restrict the growth of foci and spheroids
- 2012
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Ingår i: Histochemistry and Cell Biology. - : Springer Science and Business Media LLC. - 0948-6143 .- 1432-119X. ; 138:6, s. 881-894
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Tidskriftsartikel (refereegranskat)abstract
- Extracellular matrices (ECM) have important roles for tissue architecture, both as structural and signaling components. Members of the integrin family are the main regulators of ECM assembly and transmitters of signals from the ECM to cells. In this study, we have analyzed the role of integrin subunit beta 1 in two-dimensional (2D) and three-dimensional (3D) cell cultures using integrin beta 1 null cells (MEF beta 1(-/-) and GD25) and their beta 1 integrin-expressing counterparts. GD25 and GD25 beta 1 cells proliferated with similar kinetics in sub-confluent 2D cultures, whereas GD25 cells attained higher cell numbers in confluent culture and formed foci with fivefold higher frequency than GD25 beta 1 cells. Fibronectin fibrils were abundantly deposited throughout the GD25 beta 1 colonies but strictly limited to the central multilayered area (focus) of GD25 colonies. During 3D growth as spheroids, GD25 continuously increased in size for > 21 days while the growth of GD25 beta 1 spheroids ceased after 14 days. Similarly, MEF beta 1(-/-) cells formed foci and grew as spheroids, while the beta 1 integrin-expressing MEF did not. Expression levels of the cell cycle markers Ki67, PCNA, and histone H3-pSer10 were similar between GD25 beta 1 and GD25 spheroids. Apoptotic cells accumulated earlier in GD25 spheroids; however, cell death increased with spheroid volumes in both spheroid types. In both cell systems, the presence of beta 1 integrins resulted in higher levels of active myosin light chain and inactive myosin light chain phosphatase, and a more compact spheroid structure. In conclusion, our results reveal that regulation of 3D growth in spheroids and foci is dependent on the beta 1 subfamily of integrins, and suggest that myosin-based spheroid contraction in combination with cell death limits the growth of beta 1-expressing spheroids.
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| 2. |
- Gupta, Rajesh Kumar, et al.
(författare)
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Fibronectin Assembly in the Crypts of Cytokinesis-Blocked Multilobular Cells Promotes Anchorage-Independent Growth
- 2013
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:8, s. e72933-
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Tidskriftsartikel (refereegranskat)abstract
- Anchorage-independent growth is a characteristic feature of cancer cells. However, it is unclear whether it represents a cause or a consequence of tumorigenesis. For normal cells, integrin-mediated adhesion is required for completion of the G1 and cytokinesis stages of the cell cycle. This study identified a mechanism that can drive anchorage-independent growth if the G1 checkpoint is suppressed. Cells with defective G1 checkpoint progressed through several rounds of the cell cycle in suspension in spite of uncompleted cytokinesis, thereby forming bi- and multilobular cells. Aurora B and CEP55 were localized to midbodies between the lobes, suggesting that the cytokinesis process reached close to abscission. Integrin-mediated re-attachment of such cells induced cytokinesis completion uncoupled from karyokinesis in most cells. However, a portion of the cells instead lost the constriction and became binucleated. Also, long-term suspension culture in soft agar produced colonies where the cytokinesis block was overcome. This process was fibronectin-dependent since fibronectin-deficient cells did not form colonies unless fibronectin was expressed or exogenously added. While fibronectin normally is not deposited on non-adherent single cells, bi/multilobular cells accumulated fibronectin in the intussusceptions. Based on our data we conclude: 1) Suppression of the G1 checkpoint allows multiple rounds of the cell cycle in detached cells and thereby enables matrix formation on their surface. 2) Uncompleted cytokinesis due to cell detachment resumes if integrin interactions are re-formed, allowing colony formation in soft agar 3) Such delayed cell division can generate binucleated cells, a feature known to cause chromosomal instability.
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| 3. |
- Wang, Xuan, et al.
(författare)
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Transcription factor ZBED6 affects gene expression, proliferation, and cell death in pancreatic beta cells
- 2013
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 110:40, s. 15997-16002
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Tidskriftsartikel (refereegranskat)abstract
- We have investigated whether the recently discovered transcription factor, zinc finger BED domain-containing protein 6 (ZBED6), is expressed in insulin-producing cells and, if so, to what extent it affects beta cell function. ZBED6 was translated from a ZC3H11A transcript in which the ZBED6-containing intron was retained. ZBED6 was present in mouse βTC-6 cells and human islets as a double nuclear band at 115/120 kDa and as a single cytoplasmic band at 95-100 kDa, which lacked N-terminal nuclear localization signals. We propose that ZBED6 supports proliferation and survival of beta cells, possibly at the expense of specialized beta cell function-i.e., insulin production-because (i) the nuclear ZBED6 were the predominant forms in rapidly proliferating βTC-6 cells, but not in human islet cells; (ii) down-regulation of ZBED6 in βTC-6 cells resulted in altered morphology, decreased proliferation, a partial S/G2 cell-cycle arrest, increased expression of beta cell-specific genes, and higher rates of apoptosis; (iii) silencing of ZBED6 in the human PANC-1 duct cell line reduced proliferation rates; and (iv) ZBED6 binding was preferentially to genes that control transcription, macromolecule biosynthesis, and apoptosis. Furthermore, it is possible that beta cells, by switching from full length to a truncated form of ZBED6, can decide the subcellular localization of ZBED6, thereby achieving differential ZBED6-mediated transcriptional regulation.
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| 4. |
- Wu, Chengjun, et al.
(författare)
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A mouse mammary epithelial cell line permissive for highly efficient human adenovirus growth
- 2013
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Ingår i: Virology. - : Elsevier BV. - 0042-6822 .- 1096-0341. ; 435:2, s. 363-371
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Tidskriftsartikel (refereegranskat)abstract
- Although a few immunocompetent animal models to study the immune response against human adenoviruses (HAdV) are available, such as Syrian hamsters and cotton rats, HAdV replication is several logs lower compared to human control cells. We have identified a non-transformed mouse epithelial cell line (NMuMG) where HAdV-2 gene expression and progeny formation was as efficient as in the highly permissive human A549 cells. HAdV from species, D and E (HAdV-37 and HAdV-4, respectively) also caused a rapid cytopathic effect in NMuMG cells, while HAdV from species A, B1, B2 and F (HAdV-12, HAdV-3, HAdV-11 and HAdV-41, respectively) failed to do so. NMuMG cells might therefore be useful in virotherapy research and the analysis of antiviral defense mechanisms and the determination of toxicity, biodistribution and specific antitumour activity of oncolytic HAdV vectors.
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