Abstract
DDX3 is an RNA chaperone of the DEAD-box family that regulates translation. Ded1, the yeast ortholog of DDX3, is a global regulator of translation, whereas DDX3 is thought to preferentially affect a subset of mRNAs. However, the set of mRNAs that are regulated by DDX3 are unknown, along with the relationship between DDX3 binding and activity. Here, we use ribosome profiling, RNA-seq, and PAR-CLIP to define the set of mRNAs that are regulated by DDX3 in human cells. We find that while DDX3 binds highly expressed mRNAs, depletion of DDX3 particularly affects the translation of a small subset of the transcriptome. We further find that DDX3 binds a site on helix 16 of the human ribosomal rRNA, placing it immediately adjacent to the mRNA entry channel. Translation changes caused by depleting DDX3 levels or expressing an inactive point mutation are different, consistent with different association of these genetic variant types with disease. Taken together, this work defines the subset of the transcriptome that is responsive to DDX3 inhibition, with relevance for basic biology and disease states where DDX3 is altered.
Highlights
Translation initiation is affected by mRNA regulatory elements
Diverse biological pathways were affected by DDX3 depletion, revealing the enrichment of histone mRNAs among the few examples of translationally upregulated transcripts, which might reflect a resistance to a widespread translation suppression rather than an increase in protein synthesis
DDX3X is an essential human gene that is altered in diverse diseases
Summary
Translation initiation is affected by mRNA regulatory elements. The DEAD-box RNA chaperone DDX3 and its yeast ortholog Ded facilitate translation initiation on mRNAs with structured 5 untranslated regions (UTRs) (1–4), a function that is essential in all eukaryotes [5]. Translation initiation is affected by mRNA regulatory elements. The DEAD-box RNA chaperone DDX3 and its yeast ortholog Ded facilitate translation initiation on mRNAs with structured 5 untranslated regions (UTRs) Previous work studied how translation is altered by DDX3 variants found in medulloblastoma [3,12], which are exclusively missense variants that preferentially target conserved residues. Hematological cancers like natural killer/Tcell lymphoma [17,18] and others [19,20] have frequent variants in DDX3X, but they are mostly truncating or frameshift variants resulting in decreased expression. Changes in gene expression occurring as a result of decreased DDX3 levels remain incompletely understood
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