Cryo-EM structure of a mammalian RNA polymerase II elongation complex inhibited by -amanitin

RNA polymerase II (Pol II) is the central enzyme that transcribes eukaryotic protein-coding genes to produce mRNA. The mushroom toxin -amanitin binds Pol II and inhibits transcription at the step of RNA chain elongation. Pol II from yeast binds -amanitin with micromolar affinity, whereas metazoan Pol II enzymes exhibit nanomolar affinities. Here, we present the high-resolution cryo-EM structure of -amanitin bound to and inhibited by its natural target, the mammalian Pol II elongation complex. The structure revealed that the toxin is located in a pocket previously identified in yeast Pol II but forms additional contacts with metazoan-specific residues, which explains why its affinity to mammalian Pol II is approximate to 3000 times higher than for yeast Pol II. Our work provides the structural basis for the inhibition of mammalian Pol II by the natural toxin -amanitin and highlights that cryo-EM is well suited to studying interactions of a small molecule with its macromolecular target.