Eukaryotic RNA synthesis is catalyzed by at least three classes of RNA Polymerases (Pol I-III). The large ribosomal RNA precursor (pre-rRNA) is transcribed by Pol I, accounting for up to 60% of total cellular RNA synthesis in Saccharomyces cerevisiae. Pre-Initiation Complex (PIC) formation is a key regulatory step in the control of gene transcription by eukaryotic RNA polymerases. We have determined the structure of Pol I transcription initiation complex at near-atomic resolution using single particle cryo-EM. The structure reveals the architecture of Core Factor binding to promoter DNA and that Pol I and promoter DNA are pre-conditioned in an elongation-competent form. We have also obtained three functional states of the Pol I Initial Transcribing Complex, which allows us to propose a molecular mechanism in which Pol I utilizes the intrinsic mobility of the DNA-bound Core Factor in the process of promoter opening. This model explains why TFIIH is not necessary for Pol I promoter opening, and implies that a similar mechanism could also be used by Pol III, the other TFIIH-independent RNA polymerase.
Eukaryotic RNA polymerase II transcription
Eukaryotic gene transcription is tightly controlled during the initiation stage, when RNA polymerase II (Pol II) and the general transcription factors (GTFs) (TFIID, TFIIA, TFIIB, TFIIE, TFIIF and TFIIH) assemble at the promoter into a PIC. The initial closed promoter complex (CC) must transition into an open complex (OC), where the melted single-stranded template DNA is inserted into the active site and Pol II locates the transcription start site (TSS). This transient OC is then converted into an initial transcribing complex (ITC), where the first phosphodiester bond forms and messenger RNA starts to be synthesized. Following abortive cycles of synthesis of short RNA products, Pol II eventually clears the promoter and a stable elongation complex (EC) forms. Our cryo-EM study, which provides nearly complete pseudo-atomic models of all the structural elements within a functional human PIC, has allowed us to define the structural transitions through the processes of DNA engagement by the CC, opening of the transcription bubble in the OC, and initiation of transcription in the ITC. The present structures constitute a comprehensive structural framework for past and future studies of the complex process of eukaryotic transcription initiation.