Understanding the roles of Arabidopsis DXO1 in mRNA capping and transcriptional pausing

Regulation of gene expression is fundamental to biology. Studies into eukaryotic gene regulation have emphasized the events leading to assembly of the transcription initiation complex of RNA polymerase II (Pol II), such as the roles of transcription factors in promoting and inhibiting this assembly. However, recent evidence has increasingly demonstrated the importance of additional layers of transcriptional regulation. The 5′ end of eukaryotic mRNAs typically contain a methylated guanosine (m7G) cap that plays essential roles in all stages of transcription and in translational initiation. The mRNA cap can be differentially methylated in response to signals that mediate gene expression. Pausing of Pol II just downstream of a promoter after formation of a short nascent transcript (promoter-proximal pausing) has been recognized as a rate limiting step universally experienced by Pol II, and is particularly important for transcriptional control of signal-responsive genes. RNA capping and Pol II pausing interplay to mediate transcriptional processes. However, there is dearth of knowledge surrounding mRNA capping, cap structures, and Pol II pausing in plants.

The Arabidopsis DXO family protein DXO1 was found by my group and others to act as a decapping enzyme for RNAs containing the non-canonical NAD cap (NAD-RNAs). In addition to its NAD-RNA decapping activity, DXO1 has other functions and its mutation leads to pleiotropic phenotypes. Our recent findings indicate that DXO1, through its plant-specific N-terminal region, interacts with proteins involved in m7G capping and Pol II pausing. In this proposal, we will use functional genomics, molecular genetics, and molecular biochemistry to reveal the RNA cap structures in Arabidopsis and the roles of DXO1 and its interacting proteins in formation of these mRNA cap structures and in Pol II pausing. This study will provide novel insights into the mechanism of transcriptional control mediated by the RNA capping/decapping machinery and its interconnection with Pol II pausing.