Poster abstracts
Poster number 45 submitted by Leanne Kelley
Defining the spatiotemporal impacts of DEAD-box helicases on RNA editing
Leanne H. Kelley (Indiana University), Suba Rajendren (Indiana University), Heather A. Hundley (Indiana University)
Abstract:
RNA binding proteins (RBPs) are crucial for proper gene expression in all organisms and the coordinated roles of these RBPs with each other and target RNAs facilitate complex regulatory mechanisms. One RBP is the RNA editing enzyme, ADAR, which is responsible for the deamination of adenosine (A) to inosine (I) on doubled-stranded regions of mRNA. ADAR editing and binding can influence gene expression, adding another layer of regulation to the transcriptome. However, ADAR levels themselves are often not altered, suggesting that other regulators of RNA editing are critical to understanding RNA editing.
In Caenorhabditis elegans, ADR-2 is the sole A-to-I editing enzyme. To identify RBPs that interact with ADR-2, immunoprecipitation followed by mass spectrometry was performed. Of the ~40 proteins that associate with ADR-2, 4 are DEAD-box helicases, including GLH-1. The ADR-2/GLH-1 interaction has been validated via co-immunoprecipitation (co-IP) experiments. Experiments will be performed to determine the effect of GLH-1 on ADR-2 RNA binding and editing. Our lab has recently generated an ADR-2 binding deficient mutant in which the highly conserved lysine residues (KKxxK) of the dsRBD are converted to EAxxA. RNA immunoprecipitation experiments with the ADR-2 EAxxA mutant show significantly diminished binding to known targets. Future experiments will use the ADR-2 EAxxA mutant to test if the ADR-2/GLH-1 interaction is dependent on ADR-2 binding to RNA.
I will also determine the impact of GLH-1 on ADR-2 binding and editing. An RNA-seq experiment with a glh-1 deletion mutant, which lacks the DEAD-box helicase domain, shows that loss of glh-1 helicase activity can have a promotive or inhibitory effect on RNA editing. Overall, these studies on the ADR-2 and GLH-1 relationship will aid in our understanding of RNA editing dynamics in regulating gene expression.
Keywords: RNA editing, C elegans, helicase