NF-IL6 represses early gene expression of human papillomavirus type 16 through binding to the noncoding region

Abstract

The expression of human papillomavirus type 16 (HPV16) early genes, including E6 and E7 transforming genes, is regulated by several cellular factors binding to the noncoding region (NCR), such as the glucocorticoid receptor, NF-I, and AP1, all of which are positive regulators. We demonstrated that the nuclear factor for interleukin 6 expression (NF-EL6) specifically binds to the HPV16 NCR ranging from nucleotides 7007 to 7766 and represses the early gene expression of HPV16. The responsive element in HPV16 NCR was determined within the region ranging from nucleotides 7454 to 7766. In this region, many binding sites for other cellular transactivators, such as NF-I and AP1, have been detected. Interestingly, three of seven binding sites for NF-I and two of two binding sites for AP1 in this region overlap with the putative NF-IL6 binding sites identified by computer analysis. Competition experiments with the oligonucleotides containing such NF-I or API sites indicated that NF-IL6 certainly binds to them. Furthermore, in a chloramphenicol acetyltransferase assay using mutant NF-IL6 expression vectors, the DNA binding domain of NF-IL6 was shown to be necessary for repression, whereas the functional domain was not. These findings indicate that repression may be caused by competition with other transcriptional activators, such as NF-I and AP1. Thus, NF-IL6 may play a significant role in the regulation of viral transcnption as a part of the host's resistance to viral infection. Human papillomavirus type 16 (HPV16) is predominant among various types of HPVs identified in genital lesions, and it has been shown to be strongly associated with the development of cervical cancers (10, 45, 54). Genetic analyses of HPV16 have revealed that it has two transforming genes (E6 and E7) which are intact and actively transcribed in cervical cancers and in the cell lines derived from cervical cancers (2, 18, 29, 34, 39-41, 56, 57). E6 and E7 proteins have also been shown to bind to the p53 gene product and retinoblastoma gene product, respectively, leading to loss of the normal function of these suppressor gene products (11, 30, 38, 43, 49). These observations strongly suggest that the E6 and E7 genes play important roles in the carcinogenesis of the uterine cervix.

Transcriptional regulation of human papillomavirus type 16 LCR by different C/EBP? isoforms

“…Our results are different from those presented in two reports studying the HPV 11 LCR in which C/ EBPb seems to act as a repressor in primary human keratinocytes [40,41]. In contrast to our results, Kyo and coworkers [31] observed that C/EBPb repressed the HPV16 LCR in several cell lines in transient transfection experiments. In this latter study as well as in several other studies reporting a repression instead of an activation of the transcriptional activity by C/EBPb, the CMV±C/EBPb I expression vector was used [17,31,42].…”

“…In contrast to our results, Kyo and coworkers [31] observed that C/EBPb repressed the HPV16 LCR in several cell lines in transient transfection experiments. In this latter study as well as in several other studies reporting a repression instead of an activation of the transcriptional activity by C/EBPb, the CMV±C/EBPb I expression vector was used [17,31,42]. CMV±C/EBPb I contains the full-length C/EBPb-gene that can produce LAP and LIP by a leaky ribosome scanning mechanism [10] ( Figure 6, lane 3 in both panels).…”

“…From this study, we conclude that C/EBPb acts as an activator on the HPV16 LCR. This suggest, that LIP, the third C/EBPb isoform, induced by IL-1 (see Figure 7A), may be involved in the repression of the HPV16 LCR activity by IL-1 [31]. Other studies have also reported that IL-1 is a strong inducer of different C/EBPb isoforms.…”

“…The majority of these binding sites were only identi®ed on the basis of sequence homologies to the C/EBPb consensus sequence obtained from the IL6 promoter [7]. DNase I±footprint experiments showed that eight binding sites were actually capable of binding puri®ed C/EBPb [31]. One report showed that C/ EBPb appeared to repress the early gene expression of HPV16 through binding to the LCR [31], suggesting that C/EBPb plays a role in the regulation of viral transcription as a part of the host's resistance to viral infection.…”

Immunology of HPV infection and HPV‐associated tumors

“…19 The cytokines exerting inhibitory effects on HPV gene expression and decreasing cell proliferation are tumor necrosis factor-alpha (TNFα), interleukin-1 (IL-1), transforming growth factor-beta (TGFβ), interferons (IFNs), and others (leukoregulin, epidermal growth factor (EGF) ( Table 2). [20][21][22][23][24][25][26][27][28][29] The mechanism of downregulation of the expression of HPV E6 and E7 oncogenes seems to involve the activation of certain transcriptional factors, 24 whereas the inhibition of cell proliferation may be primarily related to the decrease in the expression of both viral and cellular oncogenes. 20 Some cytokines (e.g.…”