Structure of cytoplasmic RNA polymerase II

RNA polymerase II (Pol II) must be assembled in the cytoplasm before it enters the nucleus, where it transcribes protein-coding genes. Although transcription by Pol II is intensively studied, how this central multi-subunit enzyme is made and the role of dedicated factors remains unclear. Here, we report the integrative structural analysis of a native human Pol II from the cytoplasm captured near the end of biogenesis. The complex contained Gdown1 and three biogenesis factors - RPAP2 and the critical small GTPases GPN1 and GPN3. Cryo-EM analysis of the complex revealed how Gdown1 and RPAP2 associate with Pol II and prevent the premature association of transcription factors. Further biochemical and cryo-EM analysis revealed how RPAP2 recruits GPN1-GPN3 to the complex, and how the assembly of the RPAP2-GPN1-GPN3 complex is controlled by GTP hydrolysis. The combined results uncover a network of interactions that chaperone cytoplasmic Pol II to prevent aberrant interactions, reveal a GTP-controlled switch during the final stages of Pol II biogenesis, and suggest a general mechanism for the action of GPN-loop GTPase family of enzymes.