RT @SternbergLab: New pre-print(s) from the Sternberg Lab in collaboration with @LeifuChangLab! We uncover an unprecedented molecular mecha….

@SternbergLab Fun collaboration with @SternbergLab, and huge thanks and congrats to Renjian, Florian, Dan, Tanner, Adriana, and the whole team!.

@SternbergLab Collectively, this work provides high-resolution insights into an unexpected mechanism of RNA-guided transcription, expanding our understanding of bacterial gene regulation and opening new avenues for programmable transcriptional control.

@SternbergLab The σ2 domain still melts the −10 region, but with minimal sequence specificity. The melted −10 bases are held by base stacking, instead of the typical recognition pocket.

@SternbergLab Rather than following the classical paradigm of σE-dependent promoter recognition, recognition of the −35 element is largely supplanted by CRISPR-Cas targeting. The σ4 domain now bridges dCas12f-gRNA to RNAP instead of reading the −35 element.

@SternbergLab The σE CTD, which is unique to dCas12f-associated σE and absent in other σE paralogs, binds directly to the RuvC domain of dCas12f.1, complementing its partially truncated structure.

@SternbergLab Fta dCas12f assembles as an asymmetric dimer with a single gRNA, similar to canonical Cas12f effectors. Formation of a 14-bp full R-loop triggers conformational changes of the lid motif in the RuvC domain to recruit σE-RNAP.

@SternbergLab We captured multiple conformational and compositional states, including the DNA-bound dCas12f-σE-RNAP holoenzyme complex, revealing how RNA-guided DNA binding leads to σE-RNAP recruitment and nascent mRNA synthesis at a precisely defined distance downstream of the R-loop.

See the biology preprint from @SternbergLab.

Our new preprint is live! In collaboration with @SternbergLab, we determined the cryo-EM structure of a dCas12f-σE-RNAP complex from F. taeanensis, revealing an RNA-guided transcription mechanism that bypasses canonical promoter motifs.