Adenovirus type 12 DNA sequences in primary hamster tumors

Following subcutaneous inoculation of newborn Wistar-Furth rats with human adenovirus type 9 (Ad9), 16 of 16 female and 0 of 11 male rats developed mammary tumors. Tumor-positive animals usually developed tumors in multiple glands. Histopathological analyses indicated that three general categories of tumor could be identified. Mammary fibroadenomas were the most common tumor type encountered, but phyllodeslike tumors and solid sarcomas were also frequently found. In situ hybridization and immunohistochemical techniques established that benign fibroadenomas were derived from mammary fibroblasts (collagen type I-and vimentin-positive cells) and that malignant tumors were derived from myoepithelial cells (collagen type IV-, vimentin-, and muscle-specific actin-positive cells). The fact that mammary tumors were limited to female rats suggested that female hormones are essential for tumor growth and development. In this regard, ovariectomy of Ad9-infected female rats prevented tumor development, while subsequent diethylstilbestrol (DES) treatment elicited tumor formation. In addition, Ad9-infected and castrated male rats which received DES also developed mammary tumors. Established male mammary tumors regressed when DES treatment was stopped and reappeared after DES treatment was resumed. Together, these results indicate that estrogen is required for both initiation and maintenance of Ad9-induced mammary tumors. Southern blot analysis of high-molecularweight tumor DNA showed that mammary tumor cells contained single or multiple integrated copies of the entire Ad9 genome. RNase protection experiments established that estrogen receptor as well as Ad9 Ela and E4 mRNAs were expressed in mammary tumors, but Ad9 E3 and, surprisingly, Elb mRNAs were not expressed at detectable levels.

Section: Morsmentioning

confidence: 99%

Human adenovirus type 9-induced rat mammary tumors

Javier

Raska

Macdonald

et al. 1991

“…In most Ad-12-transformed cell lines, the viral genome has been reported to persist in multiple copies (5,9,14,22,32), whereas in the cell lines described here which contain the whole Ad-12 genome, there appeared to be about one copy per diploid copy of cellular DNA. One possible explanation for this result is that it may reflect the fact that we used purified virus, with a low particle-to-infectivity ratio, to produce our transformed cell lines (10,24).…”

Section: Fig 1 Analysis Of Ad-12 Sequences In Trans-mentioning

confidence: 64%

Adenovirus type 12-transformed rat embryo brain and rat liver epithelial cell lines: adenovirus type 12 genome content and viral protein expression

Paraskeva¹,

Brown²,

Dunn³

et al. 1982

Southern blotting analysis, six adenovirus type 12 (Ad-12)-transformed rat embryo brain cell lines and one Ad-12-transformed rat liver epithelial line were shown to contain all or nearly all the Ad-12 genome. Another Ad-12 rat liver epithelial cell line contained a repeating structure consisting of only the left-hand 16% of the Ad-12 genome. Three Ad-12-specified proteins (molecular weights, 52,000, 41,000, and 18,000) were found by immunoprecipitation to be common to all of these cell lines. Two rat liver epithelial lines, produced from an Ad-12infected culture and previously shown to be T-antigen negative by immunofluorescence, contained no detectable Ad-12 genome or Ad-12-specified proteins. Although some of the rat embryo brain transformants had been shown previously to express "late" Ad-12 mRNA, no Ad-12 structural proteins were found to be produced by these cell lines.

“…They did not report tumor induction with the transformed cells. Lee and Mak (18) analyzed the viral genome present in primary hamster tumor cells and showed the absence of the right-end sequence of Adl2 DNA in tumor cell DNA. Data in these reports contradict the present results, in which expression of the E4 gene is required for cell transformation with the El gene.…”

Section: Discussionmentioning

confidence: 99%

Expression of the E4 gene is required for establishment of soft-agar colony-forming rat cell lines transformed by the adenovirus 12 E1 gene

Shiroki¹,

Hashimoto²,

Saito³

et al. 1984

Rat 3Y1 cells were transfected with recombinant gARC (pSV2gpt carrying the adenovirus 12 early region 1 [El] gene), and focus formation was observed in monolayer cultures after culture of cells in gpt-selective medium (Eagle medium containing 10% fetal calf serum, xanthine, thymidine, aminopterin, and mycophenolic acid) for 10 days, followed by focus formation. Transformed ElY cell lines were then established from these foci. The ElY cells were transformed morphologically similarly to cells transformed with intact adenovirus 12 DNA but formed no colonies in soft-agar culture and induced tumors in transplanted rats only after a long incubation period. For the establishment of completely transformed cells, 3Y1 cells were transformed with combinations of gARC, pE3 (pBR322 carrying the adenovirus 12 E3 gene), and gE4 (pSV2gpt carrying the adenovirus 12 E4 gene) DNA. E1-3Y cells (3Y1 cells transformed with gARC and pE3 DNA), E1-4Y cells (3Y1 cells transformed with gARC and gE4 DNA), and E1-3-4Y cells (3Y1 cells transformed with gARC, pE3, and gE4 DNA) were established. These transformed cell lines were compared for growth in Eagle medium with 2 or 10% fetal calf serum, colony formation in soft-agar culture, and tumor growth in rats transplanted with the transformed cells. Several transformed cell lines of E1-4Y and E1-3-4Y cells showed colony formation in soft-agar culture and abundant expression of the E1B gene. T antigen f was seen by immunofluorescence as flecks in these cells, in which the E4 gene was transcribed, but was not seen in ElY cells, suggesting that T antigen f was encoded by the E4 gene. The suggestion was confirmed by the observation that T antigen f was detected in COS-1 cells transfected singly with gE4 DNA by immunofluorescence with polyclonal and monoclonal antibodies. Transcription of the E4 gene was confirmed in gE4-transfected COS-1 cells. T antigen f, one of the E4 gene products, was identified as a polypeptide of molecular weight 11,000 (E4-11K) by immunoprecipitation with monoclonal antibodies. The above results also suggest that expression of the E4 gene gives cells the advantage of forming colonies in soft-agar culture. A tendency was noticed for ElB gene expression to be enhanced by E4 gene expression. The relationship between enhancement of colony formation in soft-agar culture and enhancement of E1B gene expression is discussed. Recently, evidence has accumulated showing that the transforming genes of human adenovirus are located in only a small region of the left end of the viral genome (5, 10, 29, 32, 34, 37, 39, 40). The transforming genes of the highly oncogenic adenovirus 12 (Adl2) are located in the left end of Adl2 DNA (early region 1A [ElA] and E1B). Rat cells transformed by the EcoRI-C (0 through 16.4 map units) (CY cells), HindIII-G (0 through 6.8 map units) (GY cells), and AccI-H fragments (0 through 4.7 map units) (HY cells) were established (34, 37, 40). CY and GY cells showed the phenotype of completely transformed cells, showing foci in culture, forming large colonies in ...