Correlation of DNA content, p53, T antigen, and V antigen in simian virus 40‐infected human diploid cells

Abstract

Human diploid fibroblasts (HDF) have a finite life span in cell culture which can be extended when transformed with simian virus 40 (SV40). Flow cytometric analysis of SV40-HDF transformation allowed DNA content changes to be correlated with the appearance, quantity, and distribution of T antigen, p53, and V antigen, three proteins associated with this process. These studies demonstrated a shift in the DNA content to tetraploidy, which was correlated with the age of the SV4O-HDF but not the time of infection. A significant increase of the epitope recognized by PAbl22 to host p53 and the epitope PAblOl to SV40 T antigen occurred at the same time the tetraploid population appeared. However, an antigen reactive with SV40 V antibody was present at high levels in most of the population early after infection, but the levels declined with time. The percentage of PAblOl-T antigen-positive cells increased more rapidly in cells infected at a late passage, and this was concomitant with the shift in DNA content to tetraploid. Analysis of the mean fluorescence of total, gated populations (GI, G2, and > Gt) demonstrated that a threshold level of p53 and T antigen was reached in each compartment of the cell cycle. As the transformed phenotype appeared, a population of cells was continually released into the supernatant, and although these cells had a DNA pattern similar to the monolayer cells, the T antigen and p53 levels were 3-5 times higher in the tetraploid 6 2 cells.These studies correlated the expression of proteins associated with viral transformation in HDF which vary with time and shift in DNA content.

Distinct patterns of MCM protein binding in nuclei of S phase and rereplicating SV40‐infected monkey kidney cells

“…SV40-infection of normal human fibroblasts has also been shown to result in the appearance of DI>2.0 cells shortly after infection (53) and the later emergence of a tetraploid population (54). However, the chromatin/matrix binding of MCM proteins in SV40-infected human cells has not yet been analyzed.…”

Applications of Flow Cytometry to Clinical Microbiology

“…By means of multiparametric analysis, using PI for the measurement of cellular DNA and FITC-labeled antibody for the detection of viral antigens, Lehman et al (171,179) related high levels of SV40 T antigen to the appearance of cells with tetraploid DNA content due to a cell cycle block at G 2 /M.…”

Dna ploidy status and DNA content instability within single tumors in renal cell carcinoma

“…This clonal evolution of tumors has already been observed in cell lines and in transplantable tumors (2,10,11). It is particularly well documented in the transplantable prostatic tumors of the Dunning system where duplication of the genome is associated with a markedly increased growth rate and dedifferentiation, whereas subsequent chromosomal losses are associated with attainment of metastatic ability (22).…”