A Novel Subunit of Yeast RNA Polymerase III Interacts with the TFIIB-Related Domain of TFIIIB70

Abstract

There is limited information on how eukaryotic RNA polymerases (Pol) recognize their cognate preinitiation complex. We have characterized a polypeptide copurifying with yeast Pol III. This protein, C17, was found to be homologous to a mammalian protein described as a hormone receptor. Deletion of the corresponding gene, RPC17, was lethal and its regulated extinction caused a selective defect in transcription of class III genes in vivo. Two-hybrid and coimmunoprecipitation experiments indicated that C17 interacts with two Pol III subunits, one of which, C31, is important for the initiation reaction. C17 also interacted with TFIIIB70, the TFIIB-related component of TFIIIB. The interaction domain was found to be in the N-terminal, TFIIB-like half of TFIIIB70, downstream of the zinc ribbon and first imperfect repeat. Although Pol II similarly interacts with TFIIB, it is notable that C17 has no similarity to any Pol II subunit. The data indicate that C17 is a novel specific subunit of Pol III which participates together with C34 in the recruitment of Pol III by the preinitiation complex.Transcription of small genes by RNA polymerase III (Pol III) involves a multistep assembly of transcription factors into a preinitiation complex which recruits RNA Pol III (for a review, see reference 55). The multisubunit factor TFIIIC functions in promoter recognition and acts as an assembly factor to direct the binding of the initiation factor TFIIIB to an upstream gene position. Once assembled into a highly stable protein-DNA complex at Pol III promoters, TFIIIB can direct multiple rounds of transcription by Pol III in vitro in the absence of TFIIIC (28,29). TFIIIB is composed of three loosely associated polypeptides, the TATA-binding protein (TBP) (26, 30), a general transcription factor utilized for transcription by all eukaryotic and archeal RNA polymerases (21, 44), BЉ or TFIIIB90, which has no equivalent among the other general transcription factors (33,45,47), and Brf1 or TFIIIB70, which is related to the Pol II factor TFIIB (9, 13, 36). TFIIIB70 appears as a central bridging factor between the basal components of the class III transcription machinery, in a way that is much similar to the role proposed for TFIIB in the case of class II genes. TFIIIB70 is the first target of TFIIIC-DNA complex in the TFIIIB assembly process; it interacts with TBP and with TFIIIC and Pol III via their 131 and C34 subunits, respectively (10,23,34,52). Similarly, TFIIB is a target of various transcriptional activators that facilitate its incorporation into the preinitiation complex; there it binds TBP and is involved directly in Pol II-TFIIF complex recruitment through interaction with both Pol II and TFIIF (40,46). TFIIB has further been implicated in selecting the Pol II transcription start site (19,41,43). Interestingly, the N-terminal half of TFIIIB70 is structurally homologous to the full length of TFIIB. Both proteins contain at the extreme amino terminus a cysteine-rich, putative zinc-binding domain that, in the case of TFIIB, is implicat...

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The Prion Protein Has RNA Binding and Chaperoning Properties Characteristic of Nucleocapsid Protein NCp7 of HIV-1

“…Another example of a protein with both membrane-and nucleic acid-associated functions is C17. This protein is either membrane-associated, functioning in G-protein-mediated signal transduction related to the calcitonin gene-related receptor (48) or associated with RNA polymerase III for tRNA gene transcription (49). The possibility that PrP is associated with ribosomes or other cellular nucleoprotein components is presently under investigation.…”

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“…Brf contacts DNA near the site of Ty3 integration (26,42). The primary contacts between TFIIIB and pol III are mediated by Brf (17)(18)(19)21). Within TFIIIB Brf is oriented so that a conserved C-terminal region interacts with TBP.…”

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