In-cell architecture of an actively transcribing-translating expressome

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Science  31 Jul 2020:
Vol. 369, Issue 6503, pp. 554-557
DOI: 10.1126/science.abb3758

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Integrative in-cell structural biology

In bacteria, RNA polymerases can associate with ribosomes to form transcription-translation units called expressomes. Multiple models based on structural data of in vitro reconstitution assays have been proposed for how the two machineries interface with one another. Understanding this bacteria-specific coupling mechanism offers insight regarding the central dogma of molecular biology and might be leveraged for antibiotic development. O'Reilly et al. found that the NusA protein interfaces between the two complexes. The authors combined cryo–electron tomography and cross-linking mass spectrometry to produce an integrative model of the transcribing, translating expressome of Mycoplasma pneumoniae obtained entirely from in-cell data. This approach contributes to the development of in-cell structural biology.

Science, this issue p. 554


Structural biology studies performed inside cells can capture molecular machines in action within their native context. In this work, we developed an integrative in-cell structural approach using the genome-reduced human pathogen Mycoplasma pneumoniae. We combined whole-cell cross-linking mass spectrometry, cellular cryo–electron tomography, and integrative modeling to determine an in-cell architecture of a transcribing and translating expressome at subnanometer resolution. The expressome comprises RNA polymerase (RNAP), the ribosome, and the transcription elongation factors NusG and NusA. We pinpointed NusA at the interface between a NusG-bound elongating RNAP and the ribosome and propose that it can mediate transcription-translation coupling. Translation inhibition dissociated the expressome, whereas transcription inhibition stalled and rearranged it. Thus, the active expressome architecture requires both translation and transcription elongation within the cell.

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