Multiple domains of the RNA polymerase I activator hUBF interact with the TATA-binding protein complex hSL1 to mediate transcription.

Abstract

Recent evidence suggests that transcription initiation by all three eukaryotic RNA polymerases involves a complex of the TATA-binding protein (TBP) and multiple TBP-associated factors (TAFs). Here, we map the functional domains of the nucleolar HMG box protein hUBF, which binds to the human rRNA promoter and stimulates transcription by RNA polymerase I through cooperative interactions with a distinct TBP-TAF complex, hSL1. DNase I footprint analysis of mutant hUBF proteins and of a synthetic peptide of 84 amino acids reveals that HMG box 1 is necessary and sufficient for DNA sequence specificity, whereas other HMG boxes and the amino terminus modulate the binding efficiency, hUBF contains multiple activation domains that include the acidic carboxyl terminus and three HMG boxes. HMG boxes 3 and 4 and the acidic tail contribute significantly to an extended footprinting pattern in the presence of hSL1, suggestive of specific protein-protein interactions. Moreover, the inability of xUBF from Xenopus laevis to form an initiation complex with hSL1 can be overcome by hybrid proteins containing human HMG box 4 and the acidic carboxyl terminus. These results strongly suggest an important role of transcription activation domains of hUBF in mediating interactions with the TBP-TAF complex hSL1. In recent years considerable progress has been made toward elucidating the molecular mechanisms of transcription initiation. In particular, a multitude of sequence-specific transcription factors that bind to cis-acting promoter and enhancer elements have been identified and their structure and function have been analyzed (Johnson and McKnight 1989;Mitchell and Tjian 1989). Moreover, a number of components of the basal transcriptional apparatus have been implicated as targets for activation (Sawadogo and Sentenac 1990;Greenblatt 1991;Roeder 1991; Pugh and Tjian 1992). Although domains involved in transcriptional activation by sequence-specific transcription factors have been identified, their targets in the basal transcriptional machinery remained elusive.The recent identification of coactivators and TATAbinding protein-associated factors (TAFs) that are associated with the TATA-binding protein (TBP) in the TFIID fraction of the RNA polymerase II (Pol II) system focused our attention on a novel class of transcription factors. At least some of these TAFs are required for activation by sequence-specific regulatory proteins Tjian 1990, 1991;Dynlacht et al. 1991;Tanese et al. 1991;Gill and Tjian 1992). Thus, there appear to be adaptor molecules that link the basal transcriptional apparatus to the activators. Several recent reports indicate that TBP is not only involved in transcription initiation by RNA Pol II but is also necessary for transcription by RNA polymerase III (Pol III) as well as RNA polymerase I (Pol I) (Lobo et al. 1991;Margottin et al. 1991;Simmen et al. 1991;Comai et al. 1992;Cormack and Struhl 1992;Schultz et al. 1992;White et al. 1992). These studies strongly support the unifying principle that in spite of vast differenc...

Mechanism of Repression of RNA Polymerase I Transcription by the Retinoblastoma Protein

“…In previous studies, UBF has been structurally and functionally well characterized, revealing different protein domains important for DNA binding, dimerization, transactivation, and nuclear localization (22,27,30,31,40). Here we show that large parts of UBF can be removed without affecting the association with pRb and that a polypeptide encompassing HMG boxes 1 and 2 of UBF interacted with pRb almost as efficiently as did wild-type UBF.…”

“…From previous data (22) and the present results, we propose a model in which specific binding of UBF to proximal promoter elements is mostly accomplished by HMG boxes 1 and 2. Since pRb associates with this part of UBF, it prevents DNA binding and therefore inhibits transcription complex formation.…”

Multiple interactions between RNA polymerase I, TIF‐IA and TAF I subunits regulate preinitiation complex assembly at the ribosomal gene promoter

“…Preinitiation complex formation at the mammalian ribosomal RNA gene promoter is nucleated by the synergistic action of two DNA binding proteins, the HMG-box-containing upstream binding factor UBF (Jantzen et al, 1992) and the RNA polymerase I (Pol I)-specific TBP-TAF I complex TIF-IB/SL1 (Comai et al, 1992;Eberhard et al, 1993). Pol I, together with two associated initiation factors, TIF-IA and TIF-IC, is recruited to the transcription start site by specific interaction with UBF and TIF-IB/SL1 bound to the core element of the rDNA promoter (for a review, see Grummt, 1999).…”

The RNA polymerase I transcription factor UBF is a sequence-tolerant HMG-box protein that can recognize structured nucleic acids

“…UBF has now been purified and cloned from human, frog, rat and mouse (7,(13)(14)(15)(16)(17)(18)(19)(20). The UBF protein consists of an amino terminal dimerization domain (19, 21,22) followed by five repeated 80 amino acid domains (17) termed HMG-boxes due to their similarity to the DNA binding domains of High Mobility Group (HMG) proteins 1 and 2 (16), and a highly acidic carboxyl terminal tail.…”