Nuclear RNA Exosome at 3.1 Å Reveals Substrate Specificities, RNA Paths, and Allosteric Inhibition of Rrp44/Dis3

The eukaryotic RNA exosome is an essential and conserved 3′-to-5′ exoribonuclease complex that degrades or processes nearly every class of cellular RNA. The nuclear RNA exosome includes a 9-subunit non-catalytic core that binds Rrp44 (Dis3) and Rrp6 subunits to modulate their processive and distributive 3′-to-5′ exoribonuclease activities, respectively. Here we utilize an engineered RNA with two 3′ ends to obtain a crystal structure of an 11-subunit nuclear exosome bound to RNA at 3.1 Å. The structure reveals an extended RNA path to Rrp6 that penetrates into the non-catalytic core; contacts between the non-catalytic core and Rrp44, which inhibit exoribonuclease activity; and features of the Rrp44 exoribonuclease site that support its ability to degrade 3′ phosphate RNA substrates. Using reconstituted exosome complexes, we show that 3′ phosphate RNA is not a substrate for Rrp6 but is readily degraded by Rrp44 in the nuclear exosome.