RDNA transcription protein factors

The identification of the cis-acting sequences and protein factors involved in ribosomal DNA transcription strongly suggested that the network of protein-protein and protein—DNA interactions among UBF, SL1, and the promoter elements plays a major role in the assembly of a stable preinitiation complex. A number of studies have established that UBF and SL1 play a key role in this process and are necessary to direct a high level of RNA Pol I transcription in vitro. The current stepwise model of factors assembly predicts that the binding of UBF dimer to the UCE and CORE elements is a prerequisite for the recruitment of the selectivity factor SL1 to the rDNA promoter (Fig. 4, step I). Biochemical studies have indicated that two subunits of SL1, TAFi-18 and TBP, interact directly with the carboxy-terminal acidic domain of UBF (Beckmann et al., 1995 Tuan et al., 1999 step II). This finding demonstrates that the function of the carboxyl-terminal activation domain of UBF is to recruit the essential... 

Our laboratory has been involved in the identification and characterization of the transcription factors for RNA polymerase I (Pol I)-directed transcription of ribosomal RNA genes (rDNA). To this end, we initially isolated and partially purified a transcriptionally active protein complex which contains RNA polymerase I and the essential Pol I transcription factors (1). Such a fraction was obtained from whole ceU extracts (1) or from nuclear extracts (2). Subsequently, we demonstrated that a fraction obtained during chromatography of the cell extract on a heparin sepharose coliunn could prevent nonrandom transcription of cloned rat rDNA in an in vitro system (3). The major protein in this fraction exhibited characteristics of purified poly(ADP-ribose) polymerase. The present report summarizes the properties of this protein, and describes experiments showing the dramatic appearance of accurately initiated transcript in an unfiractionated whole ceU extract or nuclear extract from a tissue foUowing addition of the protein factor. 

The present studies have demonstrated that a protein factor which exhibits properties of poly(ADP-ribose) polymerase can prevent nonspecific transcription of rat rDNA in either whole ceU extracts or nuclear extracts derived from tissues. This factor is analogous to that described for RNA polymerase Il-directed transcription (7). Highly purified fractions do not require this factor for accurate transcription of rDNA (Mealey and Jacob, unpublished data) probably due to removal of DNA-nicking enzyme(s) following purification. This observation does not preclude the importance of poly(ADP-ribose) polymerase in vivo in rDNA transcription following DNA damage. Clearly, rDNA can be nicked under these conditions and the requirement for poly(ADP-ribose) polymerase to prevent binding of RNA polymerase I to these nicks will be essential. 

As mentioned above, one consequence of stalled RNA polymerase II at a DNA adduct is activation of transcription-coupled repair [27], This effect may depend on the type of polymerase, however, since the removal of some types of DNA damage is slower from RNA-polymerase I transcribed ribosomal DNA than from a nuclear gene [160], The lower level of repair in the nucleolus could also reflect the influence of other transcription factors, such as the HMG-domain protein UBF, which bind to cisplatin-mod-ified DNA [145]. When HeLa cells were exposed to cisplatin at concentrations which did not seem to affect nuclear transcription, inhibition of rDNA gene expression was associated with the redistribution of UBF, along with other factors responsible for rRNA transcription [138], These observations indicate how cisplatin might exert a combination of effects. Transcription is stopped due to titration of essential factors by the platinum-DNA adducts, and the same proteins could shield the lesions from the repair activity. 

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