Summary
Deletions are widely distributed over the genome in the most frequently occurring human cancers and are the most abundant genetic lesion found there. Deletions are highly correlated with the slow growth phenotype of mutated animal and human cells and result in chromosomal transposition when the retained ends are joined. Transpositions are only a minor source of mutation in rapidly multiplying bacteria but are a major cause of mutations in stationary bacteria. The NIH 3T3 line of mouse cells undergoes neoplastic transformation during prolonged incubation in a stationary state and expresses the slow growth phenotype on serial subculture at low density, suggesting a relation between transformation and chromosomal deletions. To further explore the relation between neoplastic transformation and the slow growth phenotype as a surrogate for deletions, two sublines of the NIH 3T3 cells with differing competence for transformation were serially subcultured in the stationary state at confluence and tested at each subculture for transformation and growth rate. Cell death in a fraction of the population and a heritable slowdown in proliferation of most of the survivors became increasingly pronounced with successive rounds of confluence. The reduction in growth rate was not proportional to the degree of transformation of the cultures, but all of the transformed cultures were slow growers at low density. All of the discrete colonies from cloning transformed cultures developed at a lower initial rate than control colonies under optimal conditions for growth, but they continued to grow at later stages, forming multilayered colonies under conditions that inhibited the further growth of the control colonies. The results suggest that prolonged incubation of NIH 3T3 cells in the stationary state results in growth-impairing deletions over a wide range of sites in the genome, but more restricted subsets of such lesions are responsible for neoplastic transformation. These findings provide dynamic, functional support in culture for the histopathological evidence that the quiescent state of cells associated with atrophy and fibrosis plays a significant role in the origin of some cancers in experimental animals and human beings.
Similar content being viewed by others
References
Aaronson, S. A.; Todaro, G. J. Basis for the acquisition of malignant potential by mouse cells cultivated in vitro. Science 162:1024–1026; 1968.
Applegate, M. L.; Moore, M. M.; Broder, C. B.; Burrell, A.; Juhn, G.; Kasweck, K. L.; Lin, P.-F.; Wadhams, A.; Hozier, J. L. Molecular dissection of mutations at the heterozygous thymidine kinase locus in mouse lymphoma cells. Proc. Natl. Acad. Sci. USA 87:51–55; 1990.
Arber, W.; Iida, S.; Jutte, H.; Caspers, P.; Meyer, J.; Hanni, C. Rear-rangements of genetic material in Escherichia coli as observed on the bacteriophage P1 plasmid. Cold Spring Harbor Symp. Quant. Biol. 43:1197–1203; 1978.
Brand, K. G. Cancer associated with asbestosis, schistosomiasis, foreign bodies, and scars. In: Becker, F. F., ed. Cancer: a comprehensive treatise. Vol. 4. 4. New York: Plenum Press; 661–692; 1982.
Brand, K. G. Solid state carcinogenesis. In: Banbury Report: non-genotoxic mechanisms in carcinogenesis. Cold Spring Harbor Laboratory 205–213; 1984.
Burek, J. E. The pathology of aging rats. Boca Raton, FL: CRC Press; 1978.
Cairns, J. The origin of human cancers. Nature 289:353–357; 1981.
Calos, M. P.; Johnsrud, L.; Miller, J. H. DNA sequence at the integrational sites of the insertion element IS1. Cell 13:411–418; 1978.
Cher, M. L.; Bova, G. S.; Moore, D. H.; Small, E. J.; Carroll, P. R.; Pin, S. S.; Epstein, J. I.; Isaacs, W. B.; Jensen, R. H. Genetic alterations in untreated metastases and androgen-independent prostate cancer detected by comparative genomic hybridization and allelotyping. Cancer Res. 56:3091–3102; 1996.
Chow, M.; Rubin, H. Irreversibility of cellular aging and neoplastic transformation: a clonal analysis. Proc. Natl. Acad. Sci. USA 93:9793–9798; 1996.
Chow, M.; Rubin, H. Evidence for cellular aging in long term confluent cultures: heritable impairment of proliferation, accumulation of age pigments and their loss in neoplastic transformation. Mech. Ageing Dev. 89:165–184; 1996.
Chow, M.; Yao, A.; Rubin, H. Cellular epigenetics: topochronology of progressive “spontaneous” transformation of cells under growth constraint. Proc. Natl. Acad. Sci. USA 91:599–603; 1994.
Clive, C.; Johnson, K. O.; Spector, J. F. S.; Batson, A. G.; Brown, M. M. M. Validation and characterization of the L5178Y/TK− mouse lymphoma mutagen system. Mutation Res. 59:61–108; 1979.
Correa, P. Human gastric carcinogenesis: a multistep and multifactorial process—First American Cancer Society Award Lecture on Cancer Epidemiology and Prevention. Cancer Res. 52:6735–7640; 1992.
Farber, E. The multistep nature of cancer development. Cancer Res. 44:4217–4223; 1984.
Fernandez, A.; Mondal, S.; Heidelberger, C. Probabalistic view of the transformation of cultured C3H 10T1/2 mouse embryo fibroblasts by 3-methylcholanthrene. Proc. Natl. Acad. Sci. USA 77:7272–7276; 1980.
Franks, L. M. Atrophy and hyperplasia in the prostate proper. J. Pathol. Bacteriol. 68:617–621; 1954.
Hozier, J.; Sawyer, J.; Clive, D.; Moore, M. Chromosome 11 aberrations in small colony L5178Y TK−/− mutants early in their clonal history. Mutation Res. 147:237–242; 1985.
Jainchill, J. L.; Aaronson, S. A.; Todaro, G. J. Murine sarcoma and leukemia viruses: assay using clonal lines of contact-inhibited mouse cells. J. Virol. 4:549–553; 1969.
Kennedy, A. R.; Fox, M.; Murphy, G.; Little, J. B. Relationship between X-ray exposure and malignant transformation in C3H 10T1/2 cells. Proc. Natl. Acad. Sci. USA 77:7262–7266; 1980.
Kinzler, K. W.; Vogelstein, B. Lessons from hereditary colorectal cancer. Cell 87:159–170; 1996.
Liber, H. L.; Yandell, D. W.; Little, J. B. A comparison of mutation induction at the tk and hprt loci in human lymphoblastoid cells; quantitative differences are due to an additional class of mutations at the autosomal tk locus. Mutat. Res. 216:9–17; 1989.
Lin, M.-C.; Mutter, G. L.; Trivijislip, P.; Boynton, K. A.; Sun, D.; Crum, C. P. Patterns of allelic loss (LOH) in vulvar squamous carcinomas and adjacent non-invasive epithelia. Amer. J. Path. 152:1313–1318; 1998.
Little, J. B.; Nagasawa, H.; Phennig, T.; Vetrous, H. Radiation-induced genomic instability: delayed mutagenic and cytogenetic effects of x-rays and alpha particles. Radiation Res. 148:299–307; 1997.
Nawroz, H.; van der Riet, P.; Hruban, R. H.; Koch, W.; Ruppert, J. M.; Sidransky, D. Allelotype of head and neck squamous cell carcinoma. Cancer Res. 54:2488–2492; 1994.
Puck, T. T. Gamma-ray mutagenesis measurement in mammalian cells. Mutation Res. 329:173–181; 1995.
Puck, T. T.; Johnson, R.; Rasmussen, S. A system for mutation measurement in mammalian cells: application to gamma-irradiation. Proc. Natl. Acad. Sci. USA 94:1218–1223; 1997.
Revell, S. H. Relationship between chromosome damage and cell death. In: (ed.) Radiation induced chromosome damage in man. New York: Allan R. Liss, Inc.; 215–233; 1983.
Rodriguez, E.; Sreekanitaiah, C.; Chaganti, R. S. K. Genetic changes in epithelial solid neoplasia. Cancer Res. 54:3398–3406; 1994.
Rubin, H. ‘Spontaneous’ transformation as aberrant epigenesis. Differentiation 53:123–137; 1993.
Rubin, H. Experimental control of neoplastic progression in cell populations: Foulds’ rules revisited. Proc. Natl. Acad. Sci. USA 91:6619–6623; 1994.
Rubin, H. Incipient and overt stages of neoplastic transformation. Proc. Natl. Acad. Sci. USA 91:12076–12080; 1994.
Rubin, H. Cell aging in vivo and in vitro. Mech. Ageing Dev. 98:1–35; 1997.
Rubin, H.; Chow, M.; Yao, A. Cellular aging, destabilization and cancer. Proc. Natl. Acad. Sci. USA 93:1825–1830; 1996.
Rubin, A. L.; Sneade-Koenig, A.; Rubin, H. High rate of diversification and reversal among subclones of neoplastically transformed NIH 3T3 clones. Proc. Natl. Acad. Sci. USA 89:4183–4186; 1992.
Rubin, H.; Xu, K. Evidence for the progressive and adaptive nature of spontaneous transformation in the NIH 3T3 cell line. Proc. Natl. Acad. Sci. USA 86:1860–1864; 1989.
Rubin, H.; Yao, A.; Chow, M. Heritable, population-wide damage to cells as the driving force of neoplastic transformation. Proc. Natl. Acad. Sci. USA 92:4843–4847; 1995.
Rubin, H.; Yao, A.; Chow, M. Neoplastic development: paradoxical relation between impaired cell growth at low population density and excessive growth at high density. Proc. Natl. Acad. Sci. USA 92:7734–7738; 1995.
Shipley, G. D.; Ham, R. G. Improved medium and culture conditions for clonal growth with minimum serum protein and for enhanced serum-free survival of Swiss 3T3 cells. In Vitro 17:656–670; 1981.
Smith, G. J.; Bell, W. N.; Grisham, J. W. Clonal analysis of the expression of multiple transformation phenotypes and tumorigenicity by morphologically transformed 10T1/2 cells. Cancer Res. 53:500–508; 1993.
Stent, G. Molecular biology of bacterial viruses. San Francisco: W. H. Freeman and Co.; 1963.
Todaro, G. J.; Green, H. Quantitative studies of the growth of mouse embryo cells n culture and their development into established lines. J. Cell Biol. 17:299–313; 1963.
Vogelstein, B.; Fearon, E. C.; Kern, S. E.; Hamilton, S. R.; Preisinger, A. C.; Nakamura, Y.; White, R. Allelotype of colorectal carcinomas. Science 4:207–211; 1989.
Willis, R. A. Pathology of tumors. Crofts, NY: Appleton-Century; 1967.
Xia, F.; Amundson, S. A.; Nickoloff, J. A.; Liber, H. L. Different capacities for recombination in closely related human lymphoblastoid cell lines with different mutational responses to x-irradiation. Mol. Cell. Biol. 14:5850–5857; 1994.
Yao, A.; Rubin, A. L.; Rubin, H. Progressive state selection of cells in low serum promotes high density growth and neoplastic transformation in NIH 3T3 cells. Cancer Res. 50:5171–5176; 1990.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Chow, M., Rubin, H. Relation of the slow growth phenotype to neoplastic transformation: Possible significance for human cancer. In Vitro Cell.Dev.Biol.-Animal 35, 449–458 (1999). https://doi.org/10.1007/s11626-999-0051-3
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s11626-999-0051-3