scholarly journals Drosophila primary microRNA-8 encodes a microRNA encoded peptide acting in parallel of miR-8

2021 ◽  
Author(s):  
Audrey Montigny ◽  
Patrizia Tavormina ◽  
Carine Duboe ◽  
Hélène San Clémente ◽  
Marielle Aguilar ◽  
...  
Keyword(s):  
Regulatory Peptides ◽  
Ribosome Profiling ◽  
Small Peptides ◽  
Molecular Approaches ◽  
Genome Expression ◽  
Genome Wide ◽  
Short Orfs ◽  
Regulatory Potential ◽  
Regulatory Feedback Loop ◽  
And Function

SummaryBackgroundRecent genome-wide studies of many species reveal the existence of a myriad of RNAs differing in size, coding potential and function. Among these are the long non-coding RNAs, some of them producing functional small peptides via the translation of short ORFs. It now appears that any kind of RNA presumably has a potential to encode small peptides. Accordingly, our team recently discovered that plant primary transcripts of microRNAs (pri-miRNAs) produce small regulatory peptides (miPEPs) involved in auto-regulatory feedback loops enhancing their cognate microRNA expression which in turn controls plant development. Here we investigate whether this regulatory feedback loop is present in Drosophila melanogaster.ResultsWe perform a survey of ribosome profiling data and reveal that many pri-miRNAs exhibit ribosome translation marks. Focusing on miR-8, we show that pri-miR-8 can produce a miPEP-8. Functional assays performed in Drosophila reveal that miPEP-8 affects development when overexpressed or knocked down. Combining genetic and molecular approaches as well as genome-wide transcriptomic analyses, we show that miR-8 expression is independent of miPEP-8 activity and that miPEP-8 acts in parallel to miR-8 to regulate the expression of hundreds of genes.ConclusionTaken together, these results reveal that several Drosophila pri-miRNAs exhibit translation potential. Contrasting with the mechanism described in plants, these data shed light on the function of yet un-described pri-microRNA encoded peptides in Drosophila and their regulatory potential on genome expression.

scholarly journals Drosophila primary microRNA-8 encodes a microRNA-encoded peptide acting in parallel of miR-8

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Audrey Montigny ◽  
Patrizia Tavormina ◽  
Carine Duboe ◽  
Hélène San Clémente ◽  
Marielle Aguilar ◽  
...  
Keyword(s):  
Regulatory Peptides ◽  
Ribosome Profiling ◽  
Small Peptides ◽  
Molecular Approaches ◽  
Genome Expression ◽  
Genome Wide ◽  
Short Orfs ◽  
Regulatory Potential ◽  
Regulatory Feedback Loop ◽  
And Function

Abstract Background Recent genome-wide studies of many species reveal the existence of a myriad of RNAs differing in size, coding potential and function. Among these are the long non-coding RNAs, some of them producing functional small peptides via the translation of short ORFs. It now appears that any kind of RNA presumably has a potential to encode small peptides. Accordingly, our team recently discovered that plant primary transcripts of microRNAs (pri-miRs) produce small regulatory peptides (miPEPs) involved in auto-regulatory feedback loops enhancing their cognate microRNA expression which in turn controls plant development. Here we investigate whether this regulatory feedback loop is present in Drosophila melanogaster. Results We perform a survey of ribosome profiling data and reveal that many pri-miRNAs exhibit ribosome translation marks. Focusing on miR-8, we show that pri-miR-8 can produce a miPEP-8. Functional assays performed in Drosophila reveal that miPEP-8 affects development when overexpressed or knocked down. Combining genetic and molecular approaches as well as genome-wide transcriptomic analyses, we show that miR-8 expression is independent of miPEP-8 activity and that miPEP-8 acts in parallel to miR-8 to regulate the expression of hundreds of genes. Conclusion Taken together, these results reveal that several Drosophila pri-miRs exhibit translation potential. Contrasting with the mechanism described in plants, these data shed light on the function of yet undescribed primary-microRNA-encoded peptides in Drosophila and their regulatory potential on genome expression.

scholarly journals Genome-wide identification of Arabidopsis non-AUG-initiated upstream ORFs with evolutionarily conserved regulatory sequences that control protein expression levels

2021 ◽  
Author(s):  
Yuta Hiragori ◽  
Hiro Takahashi ◽  
Noriya Hayashi ◽  
Shun Sasaki ◽  
Kodai Nakao ◽  
...  
Keyword(s):  
Regulatory Peptides ◽  
Inhibitory Effect ◽  
Ribosome Profiling ◽  
Open Reading Frames ◽  
Regulatory Sequences ◽  
Dependent Manner ◽  
Protein Coding ◽  
Genome Wide ◽  
Evolutionarily Conserved ◽  
Upstream Open Reading Frames

Upstream open reading frames (uORFs) are short ORFs found in the 5′-UTRs of many eukaryotic transcripts and can influence the translation of protein-coding main ORFs (mORFs). Recent genome-wide ribosome profiling studies have revealed that thousands of uORFs initiate translation at non-AUG start codons. However, the physiological significance of these non-AUG uORFs has so far been demonstrated for only a few of them. It is conceivable that physiologically important non-AUG uORFs are evolutionarily conserved across species. In this study, using a combination of bioinformatics and experimental approaches, we searched the Arabidopsis genome for non-AUG-initiated uORFs with conserved sequences that control the expression of the mORF-encoded proteins. As a result, we identified four novel regulatory non-AUG uORFs. Among these, two exerted repressive effects on mORF expression in an amino acid sequence-dependent manner. These two non-AUG uORFs are likely to encode regulatory peptides that cause ribosome stalling, thereby enhancing their repressive effects. In contrast, one of the identified regulatory non-AUG uORFs promoted mORF expression by alleviating the inhibitory effect of a downstream AUG-initiated uORF. These findings provide insights into the mechanisms that enable non-AUG uORFs to play regulatory roles despite their low translation initiation efficiencies.

scholarly journals Genome-Wide Identification and Characterization of Small Peptides in Maize

2021 ◽  
Vol 12 ◽  
Author(s):  
Yan Liang ◽  
Wanchao Zhu ◽  
Sijia Chen ◽  
Jia Qian ◽  
Lin Li
Keyword(s):  
Ribosome Profiling ◽  
Open Reading Frames ◽  
Translational Efficiency ◽  
Small Peptides ◽  
Bioinformatics Pipeline ◽  
Protein Coding ◽  
Genome Wide ◽  
A Genome ◽  
Intergenic Regions ◽  
New Perspective

Small peptides (sPeptides), <100 amino acids (aa) long, are encoded by small open reading frames (sORFs) often found in the 5′ and 3′ untranslated regions (or other parts) of mRNAs, in long non-coding RNAs, or transcripts from introns and intergenic regions; various sPeptides play important roles in multiple biological processes. In this study, we conducted a comprehensive study of maize (Zea mays) sPeptides using mRNA sequencing, ribosome profiling (Ribo-seq), and mass spectrometry (MS) on six tissues (each with at least two replicates). To identify maize sORFs and sPeptides from these data, we set up a robust bioinformatics pipeline and performed a genome-wide scan. This scan uncovered 9,388 sORFs encoding peptides of 2–100 aa. These sORFs showed distinct genomic features, such as different Kozak region sequences, higher specificity of translation, and high translational efficiency, compared with the canonical protein-coding genes. Furthermore, the MS data verified 2,695 sPeptides. These sPeptides perfectly discriminated all the tissues and were highly associated with their parental genes. Interestingly, the parental genes of sPeptides were significantly enriched in multiple functional gene ontology terms related to abiotic stress and development, suggesting the potential roles of sPeptides in the regulation of their parental genes. Overall, this study lays out the guidelines for genome-wide scans of sORFs and sPeptides in plants by integrating Ribo-seq and MS data and provides a more comprehensive resource of functional sPeptides in maize and gives a new perspective on the complex biological systems of plants.

scholarly journals Ribosome profiling reveals pervasive and regulated stop codon readthrough in Drosophila melanogaster

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Joshua G Dunn ◽  
Catherine K Foo ◽  
Nicolette G Belletier ◽  
Elizabeth R Gavis ◽  
Jonathan S Weissman
Keyword(s):  
Drosophila Melanogaster ◽  
Stop Codon ◽  
Ribosome Profiling ◽  
Terminal Protein ◽  
Regulate Gene Expression ◽  
Genome Wide ◽  
Nascent Peptide ◽  
Show Evidence ◽  
And Function ◽  
Stop Codon Readthrough

Ribosomes can read through stop codons in a regulated manner, elongating rather than terminating the nascent peptide. Stop codon readthrough is essential to diverse viruses, and phylogenetically predicted to occur in a few hundred genes in Drosophila melanogaster, but the importance of regulated readthrough in eukaryotes remains largely unexplored. Here, we present a ribosome profiling assay (deep sequencing of ribosome-protected mRNA fragments) for Drosophila melanogaster, and provide the first genome-wide experimental analysis of readthrough. Readthrough is far more pervasive than expected: the vast majority of readthrough events evolved within D. melanogaster and were not predicted phylogenetically. The resulting C-terminal protein extensions show evidence of selection, contain functional subcellular localization signals, and their readthrough is regulated, arguing for their importance. We further demonstrate that readthrough occurs in yeast and humans. Readthrough thus provides general mechanisms both to regulate gene expression and function, and to add plasticity to the proteome during evolution.

scholarly journals Selectivity of mRNA degradation by autophagy in yeast

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Shiho Makino ◽  
Tomoko Kawamata ◽  
Shintaro Iwasaki ◽  
Yoshinori Ohsumi
Keyword(s):  
Ribosomal Proteins ◽  
Rna Degradation ◽  
Mrna Degradation ◽  
Ribosome Profiling ◽  
Genome Wide ◽  
Tightly Coupled ◽  
Response To Stress ◽  
Transcriptional Gene Regulation ◽  
Synthesis And Degradation

AbstractSynthesis and degradation of cellular constituents must be balanced to maintain cellular homeostasis, especially during adaptation to environmental stress. The role of autophagy in the degradation of proteins and organelles is well-characterized. However, autophagy-mediated RNA degradation in response to stress and the potential preference of specific RNAs to undergo autophagy-mediated degradation have not been examined. In this study, we demonstrate selective mRNA degradation by rapamycin-induced autophagy in yeast. Profiling of mRNAs from the vacuole reveals that subsets of mRNAs, such as those encoding amino acid biosynthesis and ribosomal proteins, are preferentially delivered to the vacuole by autophagy for degradation. We also reveal that autophagy-mediated mRNA degradation is tightly coupled with translation by ribosomes. Genome-wide ribosome profiling suggested a high correspondence between ribosome association and targeting to the vacuole. We propose that autophagy-mediated mRNA degradation is a unique and previously-unappreciated function of autophagy that affords post-transcriptional gene regulation.

scholarly journals Humans and other commonly used model organisms are resistant to cycloheximide-mediated biases in ribosome profiling experiments

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Puneet Sharma ◽  
Jie Wu ◽  
Benedikt S. Nilges ◽  
Sebastian A. Leidel
Keyword(s):  
Human Cells ◽  
Ribosome Profiling ◽  
Model Organisms ◽  
Treatment Conditions ◽  
Genome Wide ◽  
Optimized Protocol ◽  
Gene Level ◽  
Translation Inhibitor ◽  
Species Specific ◽  
Conformational Restrictions

AbstractRibosome profiling measures genome-wide translation dynamics at sub-codon resolution. Cycloheximide (CHX), a widely used translation inhibitor to arrest ribosomes in these experiments, has been shown to induce biases in yeast, questioning its use. However, whether such biases are present in datasets of other organisms including humans is unknown. Here we compare different CHX-treatment conditions in human cells and yeast in parallel experiments using an optimized protocol. We find that human ribosomes are not susceptible to conformational restrictions by CHX, nor does it distort gene-level measurements of ribosome occupancy, measured decoding speed or the translational ramp. Furthermore, CHX-induced codon-specific biases on ribosome occupancy are not detectable in human cells or other model organisms. This shows that reported biases of CHX are species-specific and that CHX does not affect the outcome of ribosome profiling experiments in most settings. Our findings provide a solid framework to conduct and analyze ribosome profiling experiments.

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