Transfer of the marker for morphologically transformed phenotype by isolated metaphase chromosomes in hamster cells

Abstract

IN spite of extensive investigations of the process of carcinogenesis in vitro, the basic mechanisms of the primary events are poorly understood¹. In general, it is found that the first manifestation of the transformed phenotype involves morphological alteration of the cells. This is usually followed by an ill-defined process resulting in several further changes in phenotype including the ability of the cells to grow in agar (aga⁺), and to produce tumours in animals. In part because of the variety of phenotypes observed, but also because of the lack of suitable methods of analysis, it has been difficult to examine the process of transformation at the genetic level. We have recently initiated experiments designed to use chromosome transfer techniques to attempt such a genetic analysis^2,3 and work in several laboratories has shown that single gene markers can be transferred from cell to cell at frequencies of 10⁻⁶–10⁻⁷ by means of the metaphase chromosome^4–9. Most of the initial work was done with the genes for hypoxanthine phosphoribosyl transferase and thymidine kinase, but dominant markers such as resistance to methotrexate and ouabain can also be so transferred⁹, as can the wild-type alleles for three auxotrophic markers¹⁰. In seeking to delineate the genetic elements of carcinogenesis, we have examined the ability of purified metaphase chromosomes from Chinese hamster ovary (CHO) cells to transfer the first recognisable phenotype observed during cell carcinogenesis, that is, morphological transformation. Because of the low frequency of chromosome transfer, an appropriate method for selection of cells (transferents) which have incorporated the relevant markers from the background of recipient cells is necessary. Our experiments were based on the assumption that if transformation gene(s) could be transferred to senescent cells, then they should be able to produce colonies, whereas the majority of the cells which do not receive the appropriate marker, will not form colonies. The results presented here show that senescent hamster cells can be rescued and become transformed by transfer of purified metaphase chromosomes from CHO cells; the frequency of this event is similar to that of single gene markers.