Biology subjects

Niederer, R. O.

Publications and source records attributed to Niederer, R. O..

3 recordsLinked to original sources

bioRxiv2026

Systematic Interrogation Uncovers Targetable Translational Control Elements within the CFTR 5' UTR

The 5' untranslated region (5' UTR) of mRNAs often contain features that regulate ribosome recruitment and translation. These regulatory features represent potential targets to modulate translation of therapeutically relevant genes, such as cystic fibrosis transmembrane conductance regulator (CFTR), the gene responsible for Cystic Fibrosis. However, the CFTR 5' UTR regulatory landscape is largely unknown. To comprehensively identify regulatory elements in CFTR 5' UTR that could be targeted to modulate its translation, we measured ribosome recruitment to a pool of systematically mutated CFTR 5' UTRs using Direct Analysis of Ribosome Targeting (DART). This assay allowed us to test the effect of putative 5' UTR regulatory features as well as discover novel regulatory elements in an unbiased manner. Using this data, we validated a known repressive uAUG in a fetal-lung specific CFTR 5' UTR isoform and identified a stimulatory secondary structure in the canonical CFTR 5' UTR. Using a large set of targeted mutations, we also discovered stimulatory (CFTR 5' UTR Associated Stimulator of Translation, CAST) and repressive (CFTR 5' UTR Associated Repressor of Translation, CART) sequence features in the CFTR 5' UTR that regulate translation. Further, we found that CART is present in many human 5' UTRs, enriched for membrane proteins and ion transporters, and acts broadly as a translational repressor, suggesting this motif may regulate translation of not only CFTR but related genes. Overall, our strategy successfully identified regulatory features within the CFTR 5' UTR which represent promising therapeutic targets.

molecular biology↗

bioRxiv2023

Conserved 5-methyluridine tRNA modification modulates ribosome translocation

While the centrality of post-transcriptional modifications to RNA biology has long been acknowledged, the function of the vast majority of modified sites remains to be discovered. Illustrative of this, there is not yet a discrete biological role assigned for one the most highly conserved modifications, 5-methyluridine at position 54 in tRNAs (m5U54). Here, we uncover contributions of m5U54 to both tRNA maturation and protein synthesis. Our mass spectrometry analyses demonstrate that cells lacking the enzyme that installs m5U in the T-loop (TrmA in E. coli, Trm2 in S. cerevisiae) exhibit altered tRNA modifications patterns. Furthermore, m5U54 deficient tRNAs are desensitized to small molecules that prevent translocation in vitro. This finding is consistent with our observations that, relative to wild-type cells, trm2{Delta} cell growth and transcriptome-wide gene expression are less perturbed by translocation inhibitors. Together our data suggest a model in which m5U54 acts as an important modulator of tRNA maturation and translocation of the ribosome during protein synthesis.

biochemistry↗

bioRxiv2020

Direct analysis of ribosome targeting illuminates thousand-fold regulation of translation initiation

Translational control shapes the proteome in normal and pathophysiological conditions. Current high-throughput approaches reveal large differences in mRNA-specific translation activity but cannot identify the causative mRNA features. We developed direct analysis of ribosome targeting (DART) and used it to dissect regulatory elements within 5' untranslated regions that confer thousand-fold differences in ribosome recruitment in biochemically accessible cell lysates. Using DART, we identified novel translational enhancers and silencers, determined a functional role for most alternative 5' UTR isoforms expressed in yeast, and revealed a general mode of increased translation via direct binding to a core translation factor. DART enables systematic assessment of the translational regulatory potential of 5' UTR variants, whether native or disease-associated, and will facilitate engineering of mRNAs for optimized protein production in various systems. HighlightsO_LIDART illuminates thousand-fold differences in 5' UTR-specific translation activity C_LIO_LISNPs and alternative 5' UTR isoforms affect ribosome recruitment significantly C_LIO_LIInhibitory effects of RNA structures are highly dependent on 5' UTR context C_LIO_LI5' UTR motifs bind initiation factors directly, broadly stimulating translation C_LI

molecular biology↗