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ISSN: 1753-4887
CID: 1824682

Transformation by human papillomavirus type 16 (HPV16) DNA but not HPV6b DNA is enhanced by addition of the human cytomegalovirus enhancer

Morgan DM; Pecoraro G; Rosenberg I; Defendi V
Primary human cervical epithelial cells immortalized by human papillomavirus type 16 (HPV16) DNA exhibit altered morphology and differentiation characteristic of transformation, but show a lack of transformed phenotype relative to HPV18 DNA immortalized cells in terms of anchorage-independent growth (Pecoraro, Lee, Morgan, and Defendi, 1991, Am. J. Pathol. 138, 1-8). This is completely corrected by inserting a strong heterologous enhancer derived from human cytomegalovirus DNA upstream from the HPV16 long control region. The cells immortalized by this DNA form colonies in agar comparable to those formed by HPV18 DNA immortalized cells. The enhanced transformation capability correlates with increased levels of HPV16 E6-E7 and E5 transcripts. The HPV16 DNA containing this strong enhancer also transforms C127 mouse cells with increased efficiency and strength relative to the natural HPV16 DNA, as measured by the numbers and size of the colonies in agar. The positive effects of this strong enhancer appear specific for HPVs associated with genital malignancies such as HPV16, since HPV6b DNA (primarily in benign tumors) with or without the strong cytomegalovirus enhancer is incapable of immortalizing primary human cervical epithelial cells or allowing efficient growth of C127 mouse cells in agar. These results suggest that the diminished oncogenic properties of HPV16 versus HPV18 DNA in cultured cells and in human malignancies may reside in the long control regions of these viruses and, additionally, may define another difference in the oncogenic properties of HPVs associated with benign or malignant genital neoplasia
PMID: 1322595
ISSN: 0042-6822
CID: 13513

Determination of DNA cleavage specificity by esperamicins

Lu, M; Guo, Q; Krishnan, B; Golik, J; Rosenberg, I E; Doyle, T W; Kallenbach, N R
The esperamicins are members of a class of potent antitumor antibiotics that contain stained diacetylenic ring systems capable of forming DNA-cleaving diradicals upon reaction with thiols. Here we show that the diacetylenic ring core itself determines the sequence specificity for scission of duplex DNA): esperamicin A1, and three products of hydrolysis of the glycon, esperamicins C, D, and E, are found to retain a common sequence preference. The sugar residues exert a strong influence on the cleavage efficiency, presumably by interacting nonspecifically with DNA. The presence of a branch in the DNA is found locally to inhibit scission by esperamicins, and this effect is shown to be due to the core also.
PMID: 1741964
ISSN: 0739-1102
CID: 2076102

Human papillomavirus type 6b DNA required for initiation but not maintenance of transformation of C127 mouse cells

Morgan D; Pecoraro G; Rosenberg I; Defendi V
We describe the transformation of C127 mouse fibroblasts with human papillomavirus type 6b (HPV-6b) DNA, which is associated primarily with benign tumors of the human genital tract. The major transformed phenotype of the HPV-6b-transfected cells lines, which had been G418 selected, pooled, and maintained without subsequent selection, was tumorigenicity in nude mice. We found that, unlike that reported for other HPVs or papovaviruses, the transformed phenotype was expressed after a delay, in which the cells had undergone extensive culture passages (about 20 passages or 100 generations). Interestingly, the HPV-6b DNA had become reduced or nondetectable in copy number in the cells by the time the transformed phenotype was expressed and in most of the tumors induced by the cells in nude mice, indicating that high levels of HPV-6b DNA were not required for maintenance of the transformed phenotype. Clonal cell lines gave similar results. When continued G418 selection was used to maintain high-copy-number HPV-6b DNA, the cells were tumorigenic, indicating that high levels of HPV-6b DNA did not suppress tumorigenesis. These studies suggest that HPV-6b DNA initiates transformation of C127 cells but is dispensable for expression or maintenance of the transformed phenotype. Transformation by HPV-6b DNA in vitro may provide insights into the HPV type-specific association with benign versus malignant lesions in vivo and may elucidate some of the oncogenic processes involved in tumor progression
PMID: 2154622
ISSN: 0022-538x
CID: 21432

Transformation of C127 mouse fibroblasts by human papillomavirus 16

Morgan D; Pecoraro G; Rosenberg I; Defendi V
We have compared the ability of cloned DNAs of HPV16, a human papillomavirus associated with cervical carcinoma, and BPV1, a papillomavirus inducing skin lesions in cattle, to transform murine C127 cells. Unlike BPV1, HPV16 DNA failed to induce foci when C127 cells were transfected and maintained as monolayers; HPV16-transformed C127 cells could only be detected after cotransfection with HPV16 and pSV2neo DNA, selection for resistance to G418, and assay of pooled selectants for colony growth in agar. HPV16 and BPV1 C127 cells differed in terms of the size and morphology of their colonies in agar, but not in their colony-forming efficiencies. In addition, the tumors they induced in nude mice were clearly histologically distinct, with the HPV16 C127 tumors considerably more anaplastic. The HPV16 C127 cells contained viral DNA at high copy numbers integrated at random sites in the C127 genome, while the BPV1 C127, as expected, contained episomal BPV1 DNA molecules. The high complexity of the integrated HPV16 DNA was maintained in the pooled cells grown through extended passage in vitro, in clonal lines derived from single agar colonies, in nude mouse tumors induced by the cells, and in a nude mouse tumor-derived cell line, indicating the stability of the HPV16 sequences in the cells. HPV16 transcripts in the transformed C127 cells were present in three size classes (1.5, 2, and 4 kilobases) on Northern blots. The different transformed phenotypes in the same cell line induced by two structurally similar, yet distinct viruses imply differences in the underlying transforming mechanisms and possible different virus-host cell molecular interactions
PMID: 2833348
ISSN: 0008-5472
CID: 11121