Coupling of Transcription and Translation in Archaea: Cues From the Bacterial World

Abstract

The lack of a nucleus is the defining cellular feature of bacteria and archaea. Consequently, transcription and translation are occurring in the same compartment, proceed simultaneously and likely in a coupled fashion. Recent cryo-electron microscopy (cryo-EM) and tomography data, also combined with crosslinking-mass spectrometry experiments, have uncovered detailed structural features of the coupling between a transcribing bacterial RNA polymerase (RNAP) and the trailing translating ribosome in Escherichia coli and Mycoplasma pneumoniae. Formation of this supercomplex, called expressome, is mediated by physical interactions between the RNAP-bound transcription elongation factors NusG and/or NusA and the ribosomal proteins including uS10. Based on the structural conservation of the RNAP core enzyme, the ribosome, and the universally conserved elongation factors Spt5 (NusG) and NusA, we discuss requirements and functional implications of transcription-translation coupling in archaea. We furthermore consider additional RNA-mediated and co-transcriptional processes that potentially influence expressome formation in archaea.

Keywords: Nus; NusG; RNA polymerase; Spt4/5; archaea; expressome; ribosome.

Figure 1

Structures of the bacterial expressomes. (A) At mRNA spacings separating the RNA polymerase (RNAP) active site by more than ~35 nucleotides from the ribosomal P-site, and in absence of any coupling factor, RNAP adopts a wide range of orientations relative to the ribosome (uncoupled state; compare extent of translations and rotations). (B) Addition of the coupling factor NusG (turquoise) results in the formation of a physical link between RNAP and the ribosome (through uS10, orange) and restrains the rotational freedom. The emerging mRNA transcript (magenta) aligns with the ribosomal helicase (r-proteins uS3 and uS4, dashed circle; NusG coupled state). (C) Once the ribosome approaches RNAP further (mRNA spacings less than ~34 nucleotides), the expressome adopts a conformation similar to a previously observed collided state, where NusG can no longer form a physical bridge (collided state). (D–F) Comparison of transcription factor coupled states in Escherichia coli and Mycoplasma pneumoniae. The NusG-coupled state (E) gets stabilized by NusA in a similar relative orientation [D, compare outline with model, differences are within rotational freedom indicated in panel (A)]. In contrast, the NusA-coupled state observed in M. pneumoniae requires a major change in position and orientation of RNAP (F), compare outline and model, E. coli RNAP and ribosome were docked into deposited cryo-electron tomography (cryo-ET) map (O’Reilly et al., 2020).

Figure 2

Structural criteria and cellular processes that might mediate and influence transcription-translation coupling in archaea. (A) Possible scenarios for transcription translation coupling (TTC) in archaea derived from structural insights into expressome formation in bacteria: NusG/Spt5-coupled expressome formation is a possible scenario 1 representing the archaeal equivalent to the NusG-coupled state in E. coli (Webster et al., 2020). In addition, one could also imagine NusA-coupling (scenario 2), similar to what has been observed in M. pneumoniae (O’Reilly et al., 2020) or no coupling at all (scenario 3). (B) Analysis of conserved regions in archaeal Spt5 and uS10 using ConSurf (Ashkenazy et al., 2016). About 100 archaeal Spt5 and uS10 sequences were aligned, and their conservation score projected color-coded from white (0, not conserved) to dark green or dark-red (9, highly conserved), respectively, on the surface of Pyrococcus furiosus Spt5 and Pyrococcus abyssi uS10 structure. (C) Superimposition of archaeal and bacterial Spt5/NusG and (bacteria: dark-green, PDB: 6ZTJ, E. coli; archaea: light-green, PDB: 3P8B, P. furiosus) and uS10 (bacteria: orange, PDB: 6ZTJ, E. coli; archaea: red, PDB: 6SW9, P. abyssi). Residues important for the interaction with NusG are highlighted. (D) Conservation analysis of archaeal NusA proteins. Conservation scores from white (0, not conserved) to dark-blue (0, highly conserved) were calculated based on the comparison of archaeal NusA proteins and projected on the surface of Aeropyrum pernix NusA. A superimposed model of bacterial RNA (Beuth et al., 2005) is shown in pink. (E) The E. coli NusA structure (gray, PDB: 6FLQ) overlayed with A. pernix NusA (Shibata et al., 2007) (blue, PDB: 2CXC). (F) Cartoon depiction of the archaeal RNAP highlighting the hypothesis that the S1 domain of the stalk-forming subunit Rpo7 and the archaeal NusA form a homologue of bacterial NusA. TTC in archaea may be affected by co-transcriptional processes and features depicted in (G–I), including 5'UTR length and processing (G), co-transcriptional binding of non-coding RNAs (ncRNAs) (H) and the transcription termination pathway (I).