The objective of this proposal is to analyze the cellular and biochemical changes associated with progression of the transformed phenotype in cloned populations of type 5 adenovirus (Ad5) transformed rodent cells. Since progression may involve alterations in either the expression of viral genes, cellular genes or both, experiments will be designed to distinguish between these alternative possibilities. The function of Ad5 genes in modulating expression and progression of the transformed phenotype will be studied using a series of Fischer rat embryo (CREF) clones which contain the transforming gene(s) of Ad5, E1a (0 to 4.5 map units) or Ela + E1b (0 to 11.5 map units), and CREF cells transformed by the transforming genes of a host-range mutant of Ad5 (hr1) which is cold-sensitive for initiation and maintenance of transformation. A series of Ad5 transformed rat embryo (RE) clones which vary in their phenotypic expression of the transformed state, as monitored by anchorage-independence, will also be employed to investigate the role of the viral genome in regulating progression. The tumor promoting agent 12-0-tetradecanoyl-phorbol-13-acetate (TPA), which has been shown to enhance viral transformation and expression of the transformed phenotype, will be utilized to probe the mechanism of progression. TPA studies will be aided by using clones of Ad5-transformed RE and CREF cells resistant to TPA-enhancement of anchorage-independent growth. Experiments will be designed to: (a) define the role of E1a and E1b in progression; (b) determine if progression involves an alteration in the transcription and/or processing of integrated Ad5 genes; and (c) determine if progression is associated with an increase in genetic instability. The present investigations should permit a more accurate appraisal of the role of viral genes and tumor promoters in regulating cell transformation and progression of the transformed phenotype.
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