Ribo-seQC: comprehensive analysis of cytoplasmic and organellar ribosome profiling data

Sydow, Dominique; Harnett, Dermot; Ohler, Uwe

doi:10.1101/601468

Cited by 36 publications

(37 citation statements)

References 15 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Reads aligned to rRNA or a collection of snoRNAs, tRNAs and miRNAs were discarded. The filtered reads were mapped to the hg38 version of the human genome using STAR (Dobin et al, 2013) and count matrices built as described previously (Calviello et al, 2020) using Ribo-seQC (Calviello et al, 2019). Only reads above 24 nucleotides long were included in the count.…”

Section: Pre-processing and Alignment Of Ngs Datamentioning

confidence: 99%

DDX3X and DDX3Y are redundant in protein synthesis

Venkataramanan

Wilkins

Floor

2020

Preprint

Self Cite

View full text Add to dashboard Cite

DDX3 is a DEAD-box RNA helicase that regulates translation and is encoded by the X- and Y-linked paralogs DDX3X and DDX3Y. While DDX3X is ubiquitously expressed in human tissues and essential for viability, DDX3Y is male-specific and shows lower and more variable expression than DDX3X in somatic tissues. Heterozygous genetic lesions in DDX3X mediate a class of developmental disorders called DDX3X syndrome, while loss of DDX3Y is implicated in male infertility. One possible explanation for female-bias in DDX3X syndrome is that DDX3Y encodes a polypeptide with different biochemical activity. In this study, we use ribosome profiling and in vitro translation to demonstrate that the X- and Y-linked paralogs of DDX3 play functionally redundant roles in translation. We find that transcripts that are sensitive to DDX3X depletion or mutation are rescued by complementation with DDX3Y. Our data indicate that DDX3X and DDX3Y proteins can functionally complement each other in human cells. DDX3Y is not expressed in a large fraction of the central nervous system. These findings suggest that expression differences, not differences in paralog-dependent protein synthesis, underlie the sex-bias of DDX3X-associated diseases.

show abstract

Section: Pre-processing and Alignment Of Ngs Datamentioning

confidence: 99%

DDX3X and DDX3Y are redundant in protein synthesis

Venkataramanan

Wilkins

Floor

2020

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Count matrices for Ribo-seq and RNA-seq were built using reads mapping uniquely to CDS regions of protein-coding 3 genes, using the Bioconductor packages GenomicFeatures, GenomicFiles and GenomicAlignments. Genomic and transcript regions where extracted using Ribo-seQC (Calviello et al 2019). Only reads mapping for more than 25nt were used.…”

Section: Differential Expression Analysismentioning

confidence: 99%

DDX3 depletion represses translation of mRNAs with complex 5′ UTRs

Venkataramanan

Rogowski

Wyler

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

DDX3 is an RNA chaperone of the DEAD-box family that regulates translation. Its yeast ortholog Ded1 controls the translation of nearly all mRNAs, whereas DDX3 is thought to regulate only 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 most expressed mRNAs, depletion of DDX3 affects the translation level of only a small subset of the transcriptome. We further find that DDX3 binds a site on helix 16 of the human ribosome, placing it immediately adjacent to the mRNA entry channel and translation factor eIF4B. Translation changes caused by depleting DDX3 levels or through chemical inhibition mimicking a dominant negative allele are different. Taken together, our data defines the subset of the transcriptome that is responsive to DDX3 inhibition, with relevance for basic biology and disease states where DDX3 expression is altered.

show abstract

“…We combined both types of uORFs into a single uORF category as we detect no differential impact of each uORF category on the primary ORF TE, in accordance with previous work (Heesch et al, 2019). For the visualization of P-site tracks ( Figure S3C) we used plots generated by Ribo-seQC (Calviello et al, 2019).…”

Section: Identifying Translated Open Reading Framesmentioning

confidence: 67%

“…We used STAR to align the previous datasets mapped with Tophat2 and we found Pearson correlations > 0.99 across both methods, supporting the reproducibility of the data regardless of the mapping algorithm. Data QC of all Ribo-seq libraries was performed using Ribo-seQC v1.1 (Calviello et al, 2019).…”

Section: Sequencing Data Alignmentmentioning

confidence: 99%

“…The visualized data are a merger of all replicate SHR.BN-(3L) (grey, top) and SHR.BN-(3S) (green, bottom) Ribo-seq 29-nt footprints. Plots are generated with Ribo-seQC (Calviello et al, 2019) and modified to only display the following sections of genes: (i) 25nt before and 33nt after the start codon, followed by (ii) 33nt from the middle of the CDS, and finally (iii) 33nt before and 25nt after the stop codon (Calviello et al, 2019). (D) Western blots and dot plots with quantification of band intensities for the ER stress markers IRE1-alha phosphorylation (normalized for IRE1-alpha and TOM20 expression; top) and XBP1s production (normalized for GAPDH expression; bottom).…”

Section: Quantification and Statistical Analysesmentioning

confidence: 99%

See 1 more Smart Citation

Transcontrol of cardiac mRNA translation in a protein length-dependent fashion

Witte

Ruiz-Orera

Mattioli

et al. 2020

Preprint

View full text Add to dashboard Cite

Graphical Abstract Highlights• Genetic variability impacts protein synthesis rates in a rat model for cardiac hypertrophy • A trans locus affects stoichiometric translation rates of cardiac sarcomeric proteins • This master regulator locus induces a global, protein length-dependent shift in translation • Dysregulated ribosome assembly induces half-mer formation and affects translation initiation rate ABSTRACTLittle is known about the impact of naturally occurring genetic variation on the rates with which proteins are synthesized by ribosomes. Here, we investigate how genetic influences on mRNA translational efficiency are associated with complex disease phenotypes using a panel of rat recombinant inbred lines. We identify a locus for cardiac hypertrophy that is associated with a translatome-wide and protein length-dependent shift in translational efficiency. This master regulator primarily affects the translation of very short and very long protein-coding sequences, altering the physiological stoichiometric translation rates of sarcomere proteins. Mechanistic dissection of this locus points to altered ribosome assembly, characterized by accumulation of polysome half-mers, changed ribosomal configurations and misregulation of the small nucleolar RNA SNORA48. We postulate that this locus enhances a pre-existing negative correlation between protein length and translation initiation in diseased hearts. Our work shows that a single genomic locus can trigger a complex, translation-driven molecular mechanism that contributes to phenotypic variability between individuals.

show abstract

Ribo-seQC: comprehensive analysis of cytoplasmic and organellar ribosome profiling data

Cited by 36 publications

References 15 publications

DDX3X and DDX3Y are redundant in protein synthesis

DDX3X and DDX3Y are redundant in protein synthesis

DDX3 depletion represses translation of mRNAs with complex 5′ UTRs

Transcontrol of cardiac mRNA translation in a protein length-dependent fashion

Contact Info

Product

Resources

About