Nuclear receptor subfamily 4 group A member 2 (NR4A2) is an orphan nuclear receptor that is over-expressed in cancer and promotes cell proliferation, migration, transformation, and chemoresistance. Increased expression and function of NR4A2 have been attributed to various signaling pathways, but little is known about microRNA (miRNA) regulation of NR4A2 in cancer. To investigate the posttranscriptional regulation of NR4A2, we used a 3' untranslated region (UTR) reporter screen and identified miR-34 as a putative regulator of NR4A2. By using computer predictions, we identified and confirmed an miRNA recognition element in the 3' UTR of NR4A2 that was responsible for miR-34-mediated suppression. We next demonstrated that overexpression of exogenous miR-34 or activation of the p53 pathway, which regulates endogenous miR-34 expression, decreased NR4A2 expression. Consistent with previous reports, overexpression of NR4A2 blocked the induction of p53 target genes, including mir-34a. This was a phenotypic effect, as NR4A2 overexpression could rescue cells from p53-induced inhibition of proliferation. In summary, our results are the first characterization of a cancer-related miRNA capable of regulating NR4A2 and suggest a network and possible feedback mechanism involving p53, miR-34, and NR4A2.

DDX3X is a DEAD-box RNA helicase that has been implicated in multiple aspects of RNA metabolism including translation initiation and the assembly of stress granules (SGs). Recent genomic studies have reported recurrent DDX3X mutations in numerous tumors including medulloblastoma (MB), but the physiological impact of these mutations is poorly understood. Here we show that a consistent feature of MB-associated mutations is SG hyper-assembly and concomitant translation impairment. We used CLIP-seq to obtain a comprehensive assessment of DDX3X binding targets and ribosome profiling for high-resolution assessment of global translation. Surprisingly, mutant DDX3X expression caused broad inhibition of translation that impacted DDX3X targeted and non-targeted mRNAs alike. Assessment of translation efficiency with single-cell resolution revealed that SG hyper-assembly correlated precisely with impaired global translation. SG hyper-assembly and translation impairment driven by mutant DDX3X were rescued by a genetic approach that limited SG assembly and by deletion of the N-terminal low complexity domain within DDX3X. Thus, in addition to a primary defect at the level of translation initiation caused by DDX3X mutation, SG assembly itself contributes to global translation inhibition. This work provides mechanistic insights into the consequences of cancer-related DDX3X mutations, suggesting that globally reduced translation may provide a context-dependent survival advantage that must be considered as a possible contributor to tumorigenesis.