Structural biology of RNA polymerase III: Mass spectrometry elucidates subcomplex architecture

RNA polymerases (Pol) II and III synthesize eukaryotic mRNAs and tRNAs, respectively. The crystal structure of the 12 subunit Pol 11 is known, but only limited structural information is available for the 17 subunit Pol Ill. Using mass spectrometry (MS), we correlated masses of Pol 11 complexes with the Pol 11 structure. Analysis of Pol Ill showed that the complete enzyme contains a single copy of each subunit and revealed a 15 subunit form lacking the Pol Ill-specific subcomplex C53/37. DMSO treatment dissociated the C17/25 heterodimer of Pol Ill, confirming a peripheral location as its counterpart in Pol II. Tandem MS revealed the Pol III-specific subunits C82 and C34 dissociating as a heterodimer. C11 was retained, arguing against a stable trimeric subcomplex, C53/37/11. These data suggest that Pol Ill consists of a 10 subunit Pol II-like core; the peripheral heterodimers C17/25, C53/37, and C82/34; and subunit C31, which bridges between C82/34, C17/25, and the core.