Characterization of the function of the DEAD-box RNA helicase Dbp2

In eukaryotes, there are highly coupled mechanisms that require RNA-binding proteins to facilitate gene expression. Proper RNA structure and ribonucleoprotein (RNP) complex formation are critical for gene expression. DEAD-box proteins are the largest class of RNA helicases that play fundamental roles in RNA and RNP structure remodeling. However, the precise biological role of the vast majority of the ~ 40 members in this family has not been completely described. Therefore, my research focused on characterizing the role of the DEAD-box RNA helicase Dbp2 during gene expression in S. cerevisiae. To decipher the biological roles of DEAD-box proteins, I first demonstrated that the S. cerevisiae DEAD-box protein Dbp2 is an active RNA dependent ATPase and RNA helicase that unwinds RNA duplexes in vitro. Furthermore, I found that Dbp2 associates with actively transcribing genes via RNA and functions as a co-transcriptional RNA chaperone to promote efficient assembly of the mRNA binding proteins, Yra1, Nab2, and Mex67, onto poly(A)+RNA. This assembly is critical for 3’ end processing and mRNA export. I also showed that Yra1 interacts directly with Dbp2 and inhibits its unwinding activity by reducing single-stranded RNA-binding activity. This inhibition prevents over-accumulation of Dbp2 on mRNA and stabilization of a subset of RNA Pol II transcripts. Collectively, my work shows that Dbp2 is recruited to nascent RNA to unwind aberrant structures and facilitate assembly of RNA-binding proteins, including Yra1, Nab2, and Mex67, during transcription. Yra1 then prevents further cycles of unwinding by inhibiting the ability of Dbp2 to associate with single-stranded RNA. This sequential order of events involving regulation of a DEAD-box RNA helicase is critical for efficient mRNP assembly and proper gene expression. These findings provide ideas on how DEAD-box proteins are regulated and insights on the role of DDX5, which is the human ortholog of Dbp2 and is often overexpressed in cancer cells