Abstract
In eukaryotic ribosome biogenesis, the small subunit (SSU) processome is a metastable intermediate in the assembly of the small (40S) subunit. In the SSU processome, the ribosomal RNA domains are splayed open by the intervention of assembly factors as well as U3 snoRNA. A critical step during the transition from the SSU processome to the nearly mature pre-40S particle is the removal of the U3 snoRNA to allow the formation of the central pseudoknot, a universally conserved structure which connects all domains of the subunit and contributes to its dynamic nature during translation. We previously identified the DEAH/RHA RNA helicase Dhr1 as the enzyme responsible for displacing the U3 snoRNA and the SSU processome factor Utp14 as an activator of Dhr1. Here, we have utilized biochemical and yeast genetic methods to further characterize Dhr1. We show that the N terminus as well as an internal loop within the RecA2 domain are autoinhibitory. We found that Utp14 can activate the ATPase activity of Dhr1 lacking the autoinhibitory N-terminal loop but not full-length Dhr1. We considered the possibility that Utp14 activates Dhr1 by relieving the autoinhibition of the loop within the RecA2 domain. However, our results are more consistent with Utp14 activating Dhr1 by binding to the surface of the RecA1 and RecA2 domains rather than displacing the inhibitory loop. This position of Utp14 is distinct from how G-patch proteins activate other DEXH/RHA helicases and is consistent with our previous conclusion that Utp14 is not a canonical G-patch protein.