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Title: SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts

Abstract

Mouse embryo fibroblasts (MEFs) grow slowly after cultivation from animals, however, after an extended period of cultivation, their growth accelerates. We found that SWAP-70 deficient MEFs failed to increase growth rates. They maintain normal growth rates and proliferation cycles for at least 5 years. Complementing SWAP-70 deficiency in one of these MEF clones, MEF1F2, by expressing human SWAP-70 resulted in fast growth of the cells after further cultivation for a long period. The resulting cells show a transformation phenotype, since they grow on top of each other and do not show contact inhibition. This phenotype was reverted when sanguinarine, a putative SWAP-70 inhibitor, was added. Two SWAP-70 expressing clones were examined in detail. Even after cell density became very high their cdc2 and NFκB were still activated suggesting that they do not stop growing. One of the clones formed colonies in soft agar and formed tumors in nude mice. Lately, one more clone became transformed being able to make colonies in soft agar. We maintain 4 human SWAP-70 expressing MEF1F2 cell lines. Three out of 4 clones exhibited transforming phenotypes. The mouse SWAP-70 gene also promoted transformation of MEFs. Taken together our data suggest that SWAP-70 is not a typicalmore » oncogene, but is required for spontaneous transformation of MEFs. - Highlights: • Mouse embryo fibroblasts (MEFs) lacking SWAP-70 do not cause spontaneous transform. • Adding back of SWAP-70 to SWAP-70-deficient MEFs induces spontaneous transformation. • SWAP-70 is required for spontaneous transformation of MEFs.« less

Authors:
; ; ; ;  [1]; ;  [2];  [3];  [1]
  1. Institute of Cellular and System Medicine National Health Research Institute, Zhunan Town 35053, Miaoli County, Taiwan, ROC (China)
  2. Division of Tumor and Cellular Biochemistry Department of Medical Sciences Faculty of Medicine University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki-shi, Miyazaki 889-1692 Japan (Japan)
  3. Faculty of Medicine Carl Gustav Carus, Institute of Physiological Chemistry, Dresden University of Technology, Dresden (Germany)
Publication Date:
OSTI Identifier:
22648594
Resource Type:
Journal Article
Resource Relation:
Journal Name: Experimental Cell Research; Journal Volume: 345; Journal Issue: 2; Other Information: Copyright (c) 2016 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; AGAR; CELL PROLIFERATION; COLONY FORMATION; COMPLEMENT; CULTIVATION; EMBRYOS; FIBROBLASTS; HUMAN POPULATIONS; INHIBITION; MICE; NEOPLASMS; ONCOGENES; PHENOTYPE; PLANT GROWTH; TRANSFORMATIONS

Citation Formats

Chang, Yu-Tzu, Shu, Chung-Li, Lai, Jing-Yang, Lin, Ching-Yu, Chuu, Chih-Pin, Morishita, Kazuhiro, Ichikawa, Tomonaga, Jessberger, Rolf, and Fukui, Yasuhisa, E-mail: 990412@nhri.org.tw. SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts. United States: N. p., 2016. Web. doi:10.1016/J.YEXCR.2015.06.011.
Chang, Yu-Tzu, Shu, Chung-Li, Lai, Jing-Yang, Lin, Ching-Yu, Chuu, Chih-Pin, Morishita, Kazuhiro, Ichikawa, Tomonaga, Jessberger, Rolf, & Fukui, Yasuhisa, E-mail: 990412@nhri.org.tw. SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts. United States. doi:10.1016/J.YEXCR.2015.06.011.
Chang, Yu-Tzu, Shu, Chung-Li, Lai, Jing-Yang, Lin, Ching-Yu, Chuu, Chih-Pin, Morishita, Kazuhiro, Ichikawa, Tomonaga, Jessberger, Rolf, and Fukui, Yasuhisa, E-mail: 990412@nhri.org.tw. Fri . "SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts". United States. doi:10.1016/J.YEXCR.2015.06.011.
@article{osti_22648594,
title = {SWAP-70 contributes to spontaneous transformation of mouse embryo fibroblasts},
author = {Chang, Yu-Tzu and Shu, Chung-Li and Lai, Jing-Yang and Lin, Ching-Yu and Chuu, Chih-Pin and Morishita, Kazuhiro and Ichikawa, Tomonaga and Jessberger, Rolf and Fukui, Yasuhisa, E-mail: 990412@nhri.org.tw},
abstractNote = {Mouse embryo fibroblasts (MEFs) grow slowly after cultivation from animals, however, after an extended period of cultivation, their growth accelerates. We found that SWAP-70 deficient MEFs failed to increase growth rates. They maintain normal growth rates and proliferation cycles for at least 5 years. Complementing SWAP-70 deficiency in one of these MEF clones, MEF1F2, by expressing human SWAP-70 resulted in fast growth of the cells after further cultivation for a long period. The resulting cells show a transformation phenotype, since they grow on top of each other and do not show contact inhibition. This phenotype was reverted when sanguinarine, a putative SWAP-70 inhibitor, was added. Two SWAP-70 expressing clones were examined in detail. Even after cell density became very high their cdc2 and NFκB were still activated suggesting that they do not stop growing. One of the clones formed colonies in soft agar and formed tumors in nude mice. Lately, one more clone became transformed being able to make colonies in soft agar. We maintain 4 human SWAP-70 expressing MEF1F2 cell lines. Three out of 4 clones exhibited transforming phenotypes. The mouse SWAP-70 gene also promoted transformation of MEFs. Taken together our data suggest that SWAP-70 is not a typical oncogene, but is required for spontaneous transformation of MEFs. - Highlights: • Mouse embryo fibroblasts (MEFs) lacking SWAP-70 do not cause spontaneous transform. • Adding back of SWAP-70 to SWAP-70-deficient MEFs induces spontaneous transformation. • SWAP-70 is required for spontaneous transformation of MEFs.},
doi = {10.1016/J.YEXCR.2015.06.011},
journal = {Experimental Cell Research},
number = 2,
volume = 345,
place = {United States},
year = {Fri Jul 15 00:00:00 EDT 2016},
month = {Fri Jul 15 00:00:00 EDT 2016}
}
  • The mechanism of in vitro transformation of the mouse embryo fibroblast C3H/10T 1/2 clone 8 by aflatoxin B1 (AFB1) was studied in confluent holding (CH) experiments. Confluent cultures of C3H/10T 1/2 cells were treated with AFB1 for 16 hours, and the DNA adduct composition and concentration were determined by chromatographic procedures after 0, 8, 16, and 40 hours of CH when the cells were replated at low density for the expression of their colony-forming ability and the formation of transformed foci. Total adduct concentration and the concentration of the major primary adduct 2,3-dihydro-2-(N7-guanyl)-3-hydroxyaflatoxin B1 (AFB1-N7-Gua) decreased continuously during CH duemore » to spontaneous decomposition and probably also due to enzymatic repair processes. In contrast, the more chemically stable secondary product 2,3-dihydro-2-(N5-formyl-2',5',6'-triamino-4'-oxo-N5-pyrimidyl)-3-hydroxyaflatoxin B1 (AFB1-triamino-Py) accumulated in the DNA and reached its maximum concentration after 16 hours of CH. While the loss of total AFB1-DNA adducts during CH was reflected in recovery of viability, the potential to form transformed foci reached a maximum after 16 hours of CH and then decreased with continued CH below the initial value. Therefore, no simple relationship exists between the concentration of the total adducts AFB1-N7-Gua and AFB1-triamino-Py at the time of release from CH and the potential to form transformed foci. However, DNA lesions or abnormal DNA configurations formed during CH as a consequence of the cellular processing of AFB1-DNA adducts may play a role in the transformation process.« less
  • There is much evidence from in vivo and in vitro carcinogenesis studies that active oxygen species play a role in tumor promotion. We tested directly whether superoxide produced extracellularly by xanthine-xanthine oxidase (X-XO) has the capacity to promote initiated mouse embryo C3H/10T1/2 fibroblasts. Cell cultures initiated with either /sup 137/Cs ..gamma..-rays or benzo(a)pyrene diol epoxide I were found to transform 3-30 times more effectively when subsequently treated daily for 3 weeks with nontoxic doses of X-XO. Scavengers of active oxygen radicals such as superoxide dismutase or superoxide dismutase in combination with catalase reduced the frequency of appearance of transformed focimore » by 3-25 times when compared to cultures receiving X-XO alone. These results show that active oxygen species such as superoxide and H/sub 2/O/sub 2/ can act in a promotional manner that mimics the effects of the mouse skin promoter phorbol 12-myristate 13-acetate in this system. X-XO also acted as a weak complete carcinogen. 27 references, 4 figures, 1 table.« less
  • Whole Chinese hamster embryo lineages have been shown to undergo multistep spontaneous neoplastic progression during serial passage in culture. The authors have studied the binding, internalization, and degradation of /sup 125/I-labeled epidermal growth factor at four different stages of transformation. The whole Chinese hamster embryo cells lost cell surface epidermal growth factor receptors gradually during the course of neoplastic progression until only 10% of the receptor number present in the early-passage cells (precrisis) were retained in the late-passage cells (tumorigenic). No differences in internalization rates, chloroquine sensitivity, or ability to degrade hormone between the various passage levels were seen. Nomore » evidence for the presence in conditioned medium of transforming growth factors which might mask or down-regulate epidermal growth factor receptor was obtained. These results suggest that a reduction in cell surface epidermal growth factor receptor might be an early event during spontaneous transformation in whole Chinese hamster embryo cells.« less