SNAP19 mediates the assembly of a functional core promoter complex (SNAP c ) shared by RNA polymerases II and III

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The human small nuclear RNA (snRNA) 1 genes, which encode snRNAs that are involved in RNA processing reactions such as mRNA splicing, serve as prototypes for a family of genes whose promoters are characterized by the presence of a proximal sequence element (PSE) and a distal sequence element (DSE). From a transcription point of view, this family of genes is highly interesting because all of its members have very similar promoters, even though some of them are transcribed by RNA polymerase (pol) II and others by pol III. As a result, the snRNA genes have served as a model system to explore how RNA polymerase specificity is determined and, in general, to compare the pol II and III transcription machineries. This has led to the concept that the pol II and III transcription machineries use common factors, the best known of which is the TATA box binding protein (TBP). In addition, the relative simplicity of these promoters has also made them an attractive system to study how transcriptional activators perform their function. Fig. 1 shows the structures of snRNA promoters from Homo sapiens (Hs), Arabidopsis thaliana (At), and Drosophila melanogaster (Dm) and serves to illustrate the remarkable fact that although snRNA promoters have diverged during evolution, the close similarity between those recognized by pol II and those recognized by pol III has been conserved. In fact, in each of the examples in Fig. 1, RNA polymerase specificity can be changed by altering a single parameter, indicated in red on the figure. The Structure of snRNA PromotersIn the human genes, the U1 and U2 snRNA promoters serve as the prototypic pol II snRNA promoters, and the U6 snRNA promoter serves as the prototypic pol III snRNA promoter (see Ref. 1 for a review). The human pol II snRNA core promoters contain only one essential element, the PSE, whereas the pol III snRNA core promoters consist of two elements, the PSE and a TATA box located at a fixed distance downstream. The DSE serves to enhance transcription from the core promoter. Both the DSE and the PSE can be interchanged between pol II and III snRNA promoters with no effect on RNA polymerase specificity, which is determined by the presence or absence of the TATA box. The A. thaliana pol II and III snRNA promoters contain an upstream sequence element (USE) and a TATA box, which are both interchangeable between the pol II and III snRNA promoters. RNA polymerase specificity is determined in this case by the exact spacing between the USE and the TATA box, which is 33-34 base pairs (bp) and 23-24 bp in the pol II and III snRNA promoters, respectively (2).The D. melanogaster pol II snRNA promoters contain two elements referred to as the PSEA and the PSEB spaced by 8 bp, and the pol III snRNA promoters contain a PSEA and a TATA box spaced by 12 bp. The PSEA is quite conserved in various pol II and III snRNA promoters, but positions 19 and 20 of the 21-bp elements are always g/aG in the pol II and TC in the pol III snRNA promoters. RNA polymerase specificity is determined by the precise sequ...

Section: The Pse Binding Factorsmentioning

confidence: 99%

Small Nuclear RNA Genes: a Model System to Study Fundamental Mechanisms of Transcription

Hernandez

2001

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195

201

“…U1 and U2) and RNA polymerase III (e.g. U6 and 7SK) (4,5) and purified or recombinant PTF functions as a basal transcription factor for both types of snRNA gene (6,7). The pol III-specific core promoters contain an additional TATA-box at Ϫ25, which in this context is responsible for the selective recruitment of pol III (5,8).…”

mentioning

confidence: 99%

BRFU, a TFIIB-like Factor, Is Directly Recruited to the TATA-box of Polymerase III Small Nuclear RNA Gene Promoters through Its Interaction with TATA-binding Protein

Čabart

Murphy

2001

template. We have mapped amino acid residues within TBP and domains of BRFU that mediate this interaction. BRFU has no specificity for sequences flanking the TATA-box and also forms a stable complex on the TATA-box of the pol II-specific adenovirus major late promoter (AdMLP). Furthermore, pol III-type transcription can initiate from an snRNA gene promoter containing an AdMLP TATA-box and flanking sequences. Therefore, the polymerase recruitment is not simply determined by the sequence of the TATA-box and immediate flanking sequences.The core promoter regions of human snRNA 1 genes are sufficient to direct low levels of transcription in vitro and contain a binding site, called the proximal sequence element (PSE), for the multisubunit factor PBP/PTF/SNAP c (1-3). The PSE, usually located around Ϫ55, is interchangeable between snRNA gene promoters recognized by RNA polymerase II (e.g. U1 and U2) and RNA polymerase III (e.g. U6 and 7SK) (4, 5) and purified or recombinant PTF functions as a basal transcription factor for both types of snRNA gene (6, 7). The pol III-specific core promoters contain an additional TATA-box at Ϫ25, which in this context is responsible for the selective recruitment of pol III (5,8). Insertion of a TATA element into the pol II-transcribed U2 promoter converts it into a predominantly pol III promoter (8), whereas mutation of the 7SK TATA-box reduces pol III transcription and allows snRNA-type transcription by pol II to occur (5). TBP is required for transcription of both types of snRNA gene and is likely to be recruited to the TATAless pol II-specific promoters by interaction with PTF binding to the PSE (3). PTF also potentiates direct binding of TBP to the TATA-box of the pol III-specific promoters (9). Because loss of pol III transcription correlates with the loss of TBP binding to the mutated TATA-box (5, 10), the differential interaction of TBP with template DNA and the other proteins of the PIC is likely to play a key role in the ultimate recruitment of different polymerases.For transcription of tRNA and 5 S rRNA genes, which have gene-internal pol III promoters, TBP is associated with TFIIIB90 (11) (also called hBRF (12)) and hBЉ (13) (also called TFIIIB150 (14)) within the TFIIIB-␤ complex (15). At these TATA-less promoters, the internal promoter recruits TFIIIC that results in the subsequent recruitment of TFIIIB-␤, which may then directly recruit pol III. TBP is a more loosely associated subunit of the less well characterized snRNA-specific TFIIIB form, designated hTFIIIB-␣ (15), which is required for transcription of the U6/7SK genes by pol III. Recently, a basal transcription factor known as BRFU (13), or TFIIIB50 (14), has been shown to be required for transcription of these snRNA genes but not an adenovirus 2 VA1 gene with an internal pol III promoter. Interestingly, BRFU/TFIIIB50 has sequence homology to both TFIIIB90/BRF and the pol II initiation factor TFIIB. A complex of TFIIIB50 and four tightly associated factors constitutes, together with TBP and TFIIIB150, the complete ...

“…During late G 2 /early prophase, the SNAP45 staining was still largely within regions of DAPI staining ( Fig. 1D, panels [5][6][7][8], but starting in prometaphase and up to anaphase, it was largely nonoverlapping with the DAPI staining of DNA (panels 9 -20). In telophase, the SNAP45 staining started to concentrate again in the two newly formed nuclei (Fig.…”

Section: Centrosome and Mid-body Staining With An Anti-snap45mentioning

confidence: 96%

“…SNAP190 forms the backbone of the complex and binds three of the four remaining subunits through two main regions as follows: a region within the N-terminal third binds SNAP19 and SNAP43, whereas a region close to the C terminus of the protein binds SNAP45. SNAP50 joins the complex through contacts with SNAP43 (5)(6)(7)(8). A subcomplex of SNAP c , referred to as mini-SNAP c , missing the C-terminal two-thirds of SNAP190 and associated SNAP45, is still capable of directing in vitro transcription of RNA polymerase II and III snRNA genes, albeit with lower efficiency than complete SNAP c (7).…”

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confidence: 99%

Mitotic Functions for SNAP45, a Subunit of the Small Nuclear RNA-activating Protein Complex SNAPc

Shanmugam

Hernandez

2008

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The small nuclear RNA-activating protein complex SNAP c is required for transcription of small nuclear RNA genes and binds to a proximal sequence element in their promoters. SNAP c contains five types of subunits stably associated with each other. Here we show that one of these polypeptides, SNAP45, also known as PTF ␦, localizes to centrosomes during parts of mitosis, as well as to the spindle midzone during anaphase and the mid-body during telophase. Consistent with localization to these mitotic structures, both down-and up-regulation of SNAP45 lead to a G 2 /M arrest with cells displaying abnormal mitotic structures. In contrast, down-regulation of SNAP190, another SNAP c subunit, leads to an accumulation of cells with a G 0 /G 1 DNA content. These results are consistent with the proposal that SNAP45 plays two roles in the cell, one as a subunit of the transcription factor SNAP c and another as a factor required for proper mitotic progression.Many biological processes are combinatorial, using the principle of mixing limited numbers of individual elements to give rise to nearly unlimited numbers of combinations with different functional attributes. A classical example occurs in promoters, where different arrangements of sequence elements result in the recruitment of different combinations of transcription factors that can provide the complex regulation needed for processes such as differentiation and development. Another example is in the repeated use of various polypeptides in different protein complexes. In some cases, such as the TBP-associated factors (TAFs) present in both the transcription factor IID (TFIID) and Spt-Ada-Gen5 acetyltransferase (SAGA) complexes (1, 2), the resulting complexes are involved in the same general process, in this example transcription. In other cases, however, the same proteins can exert their effect in completely different processes; for example, glyceraldehyde-3-phosphate dehydrogenase functions as a glycolytic enzyme in the cytoplasm as well as a member of a nuclear co-activator complex involved in cell cycle-regulated transcription from the H2B promoter (3). This last theme is becoming more and more common as we learn more about the players in various cellular processes.The snRNA-activating protein complex SNAP c is a multisubunit complex containing five types of subunits, SNAP190, SNAP50, SNAP45, SNAP43, and SNAP19, that is required for RNA polymerase II and III transcription of the human snRNA 2 genes (for a review see Ref. 4). The arrangement of the subunits within the complex has been deduced from protein-protein interaction studies and reconstitution of partial complexes in vitro. SNAP190 forms the backbone of the complex and binds three of the four remaining subunits through two main regions as follows: a region within the N-terminal third binds SNAP19 and SNAP43, whereas a region close to the C terminus of the protein binds SNAP45. SNAP50 joins the complex through contacts with SNAP43 (5-8). A subcomplex of SNAP c , referred to as mini-SNAP c , missing the C-terminal...