Transcriptional Initiation in Ribosomal Protein Genes in the Fission Yeast Schizosaccharomyces pombe

Transcription of class II genes in eukaryotic organisms is carried out by the multi-subunit enzyme RNA polymerase II (RNA pol II) and includes the general transcription factors and the mediator. The region inside the promoters, which recruits and specifies the transcriptional machinery, is called “core promoter” and contains sub-regions called “core promoter elements,” which are necessary for transcription initiation, where the most studied and classic element is the TATA-box. Ribosome protein gene (RPG) promoters do not possess a TATA-box (TATA-less promoters), and those, in particular, in the fission yeast Schizosaccharomyces pombe have a TATA-box analog called the HomolD-box. The transcription of RPG promoters is dependent on the RNA pol II transcription system and the HomolD-box is recognized by the transcription factor Rrn7. In this chapter, the authors will describe the general mechanisms associated to the transcription of TATA-less promoters in eukaryotic organisms and how the transcription initiation is carried out in the RPG promoters from those organisms, particularly in Schizosaccharomyces pombe. Finally, the authors will analyze the role of the HomolD-box and the transcription factor Rrn7 in the coordination of transcription initiation from RPG promoters and other ribosome-related genes and the presence of transcriptional modules in their promoters, which could be coordinated and regulated by a discrete number of transcription factors

Tobacco Exposure Enhances Human Papillomavirus 16 Oncogene Expression via EGFR/PI3K/Akt/c-Jun Signaling Pathway in Cervical Cancer Cells

High-risk human papillomavirus (HR-HPV) infection is not a sufficient condition for cervical cancer development because most infections are benign and naturally cleared. Epidemiological studies revealed that tobacco smoking is a cofactor with HR-HPV for cervical cancer initiation and progression, even though the mechanism by which tobacco smoke cooperates with HR-HPV in this malignancy is poorly understood. As HR-HPV E6/E7 oncoproteins overexpressed in cervical carcinomas colocalize with cigarette smoke components (CSC), in this study we addressed the signaling pathways involved in a potential interaction between both carcinogenic agents. Cervical cancer-derived cell lines, CaSki (HPV16; 500 copies per cell) and SiHa (HPV16; 2 copies per cell), were acutely exposed to CSC at various non-toxic concentrations and we found that E6 and E7 levels were significantly increased in a dose-dependent manner. Using a reporter construct containing the luciferase gene under the control of the full HPV16 long control region (LCR), we also found that p97 promoter activity is dependent on CSC. Non-synonymous mutations in the LCR-resident TPA (12-O-tetradecanoylphorbol 13-acetate)-response elements (TRE) had significantly decreased p97 promoter activation. Phosphoproteomic arrays and specific inhibitors revealed that CSC-mediated E6/E7 overexpression is at least in part reliant on EGFR phosphorylation. In addition, we showed that the PI3K/Akt pathway is crucial for CSC-induced E6/E7 overexpression. Finally, we demonstrated that HPV16 E6/E7 overexpression is mediated by JUN. overexpression, c-Jun phosphorylation and recruitment of this transcription factor to TRE sites in the HPV16 LCR. We conclude that acute exposure to tobacco smoke activates the transcription of HPV16 E6 and E7 oncogenes through p97 promoter activation, which involves the EGFR/PI3K/Akt/C-Jun signaling pathway activation in cervical cancer cells

Transcriptional functions of a new mammalian TATA-binding protein- related factor

A mammalian protein highly homologous to TATA-binding protein (TBP) has been identified and cloned. The recombinant mammalian TBP-related factor binds to the TATA box of the Ad-MLP and forms stable complexes with TFIIB on the promoter DNA. The mammalian TBP-related factor is able to substitute for TBP in supporting transcription by RNA polymerase II in an in vitro reconstituted system

Factors involved in specific transcription by mammalian RNA polymerase II. Factors IIE and IIF independently interact with RNA polymerase II.

The purification and characterization of transcription factor IIF (TFIIF), a factor required for transcription by the RNA polymerase II machinery, is described. TFIIF was isolated from the previously described IIE protein fraction. TFIIF enters into the transcription cycle via a preinitiation complex, and it is required for the formation of a complex capable of initiating transcription in the presence of heparin concentrations that inhibit the action of a free factor. TFIIF and TFIIE independently interacted with purified RNA polymerase II. TFIIF and TFIIE were both required for transcription of several class II promoters, including a promoter that lacks the conserved TATA box. Interestingly TFIIF was absolutely required for the formation of a preinitiation complex; however, it also affected the elongation phase of the transcription cycle. TFIIF, together with the previously described elongation factor TFIIS, was required for efficient elongation

Affinity Purification of a Human RNA Polymerase II Complex Using Monoclonal Antibodies against Transcription Factor IIF

Five different monoclonal antibodies that immunoreact with RAP74, the large subunit of general transcription factor (TF) IIF, were produced and characterized. Using one of these antibodies, an affinity purification procedure was devised to isolate a human RNA polymerase II complex. This procedure is fast, simple, and reproducible and does not require extensive purification. The RNA polymerase II complex isolated using this procedure contains SRB (suppressor of RNA polymerase B) polypeptides, transcription factors IIE and IIF, limiting amounts of TFIIH, and the TATA-binding protein, but was devoid of TFIIB