RNA polymerase II transcription complexes imaged in near-native state for first time

Researchers at Pennsylvania State University have captured the first detailed structural views of eukaryotic RNA polymerase II transcription complexes as they exist inside living cells. The team isolated intact complexes directly from fruit fly embryos, preserving associated proteins, DNA, RNA, and nucleosomes, then imaged them using cryo-electron microscopy.

Previous structural studies relied on highly purified polymerase II assembled under idealized laboratory conditions. Those samples yielded a single, uniform structure composed of 12 subunits. The new near-native samples revealed a heterogeneous population: some complexes contain all 12 subunits, while others lack two subunits and operate with only 10.

The variation was unexpected because the 12-subunit model had been considered the standard functional form. The presence of 10-subunit complexes in living embryos suggests that polymerase II composition may change during the transcription cycle or in response to cellular conditions.

Computational sorting of thousands of cryo-EM images allowed the researchers to reconstruct multiple distinct three-dimensional structures from the same sample. This approach moves beyond the traditional goal of solving a single representative structure and instead maps the structural diversity present in a living system.

The work began in 2021 when emeritus professor David Gilmour showed lead investigator Katsuhiko Murakami a crude polymerase II preparation from fruit fly embryos. Though impure by conventional standards, the sample retained the native complexity that purified systems discard, prompting the team to develop extraction and imaging methods suited to such material.

Co-author Jean-Paul Armache described the shift as moving from a sanitized, single snapshot to a messier but more accurate picture of molecular biology in action. The researchers say the method can be applied to other organisms and cellular processes, including studies in archaea, to observe molecular machines in their natural environments.

The study was published in Nature Communications. The authors note that understanding transcription in its native context provides a more accurate blueprint of cellular function, with potential implications for medicine and other fields.