Abstract
Translation efficiency can be affected by mRNA stability and secondary structures, including G-quadruplex structures (G4s). The highly conserved DEAH-box helicase DHX36/RHAU resolves G4s on DNA and RNA in vitro, however a systems-wide analysis of DHX36 targets and function is lacking. We map globally DHX36 binding to RNA in human cell lines and find it preferentially interacting with G-rich and G4-forming sequences on more than 4500 mRNAs. While DHX36 knockout (KO) results in a significant increase in target mRNA abundance, ribosome occupancy and protein output from these targets decrease, suggesting that they were rendered translationally incompetent. Considering that DHX36 targets, harboring G4s, preferentially localize in stress granules, and that DHX36 KO results in increased SG formation and protein kinase R (PKR/EIF2AK2) phosphorylation, we speculate that DHX36 is involved in resolution of rG4 induced cellular stress.
Highlights
Translation efficiency can be affected by messenger RNA (mRNA) stability and secondary structures, including G-quadruplex structures (G4s)
Loss-of-function analysis with DHX36-KO cells coupled with RNA sequencing (RNA-seq), ribosome profiling (Ribo-seq), and high throughput proteomics show that binding of DHX36 in the 3′ and 5′ untranslated regions (UTR) results in higher target mRNA translational efficiency, in a helicase activity-dependent manner
DHX36 is a cytoplasmic helicase interacting with mRNA
Summary
Translation efficiency can be affected by mRNA stability and secondary structures, including G-quadruplex structures (G4s). In vitro and in silico approaches have revealed over 13,000 sites in the human transcriptome with the potential to form rG4s8–10, further supported by immunofluorescence experiments with a specific antibody that detected rG4s in the cytoplasm of human cells[11] These rG4s may influence many aspects of posttranscriptional regulation, including alternative polyadenylation, splicing, and miRNA biogenesis[3,7,12,13,14,15]. Loss-of-function analysis with DHX36-KO cells coupled with RNA sequencing (RNA-seq), ribosome profiling (Ribo-seq), and high throughput proteomics show that binding of DHX36 in the 3′ and 5′ UTR results in higher target mRNA translational efficiency, in a helicase activity-dependent manner. We propose the model that DHX36 loss results in the formation of rG4s and other structures on target mRNAs that stabilize them, and trigger a stress response rendering them translationally incompetent, possibly by sequestration in SGs or their precursors/seeds
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