Uncultured protist
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ABSTRACT:
PROVIDER: PRJNA305513 | ENA |
REPOSITORIES: ENA
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Project description:Cleavage and polyadenylation are the final steps of eukaryotic mRNA 3' end formation. The most critical element in humans and other model organisms is the poly(A) signal, an AAUAAA hexamer. We recently discovered that the deeply branching eukaryote – Giardia lamblia uses a different but well-defined poly(A) signal, AGURAA. To better characterize when this evolutionary shift in the poly(A) signal occurred, we performed direct RNA sequencing of four protists within the Metamonada supergroup and two outgroup protists. Both outgroup protists and the non-Giardia Metamonada species use the AAUAAA poly(A) signal, indicating it is the ancestral signal. In contrast, all Giardia species use the WGURAA poly(A) signal, indicating it is a derived feature within Giardia or Fornicata. The change in this ubiquitous regulatory element raises questions about the sequence features that specify genuine poly(A) sites and how to avoid premature cleavage in the coding sequence. Therefore, we used a sequence classifier, a gapped k-mer support vector machine, that could discriminate between WGURAA sites in 3'UTRs and those in the CDS (F1 = 0.97). We found that Giardia lamblia uses nucleotides directly flanking the poly(A) signal for its recognition, with downstream nucleotides being the most important. Another member of the Giardia genus, Giardia muris, uses a different strategy: almost complete depletion of WGURAA hexamers in coding sequences. Ones that remain in the coding sequence can be recognized as poly(A) signals and undergo premature cleavage. These results identify unique features of the Giardia pathogens that could be targeted for drugs and highlight the diversity and evolution of mRNA 3' end formation in eukaryotes.
2024-08-01 | GSE260731 | GEO
Project description:Ribosome biogenesis is essential for protein synthesis in gene expression. Yeast eIF5B has been shown biochemically to facilitate 18S rRNA 3’ end maturation during late-40S ribosomal subunit assembly and gate the transition from translation initiation to elongation. But the effects of eIF5B have not been studied at the genome-wide level in any organism, and 18S rRNA 3’ end maturation is poorly understood in plants. Arabidopsis HOT3/eIF5B1 was found to promote development and heat-stress acclimation by translational regulation, but its molecular function remained unknown. Here, we show that HOT3 is a late-stage ribosome biogenesis factor that facilitates 18S rRNA 3’ end processing and is a translation initiation factor that globally impacts the transition from initiation to elongation. By developing and implementing 18S-ENDseq, we revealed previously unknown events in 18S rRNA 3’ end maturation or metabolism. We quantitatively defined new processing hotspots and identified adenylation as the prevalent non-templated RNA modification at the 3’ ends of pre-18S rRNAs. Aberrant 18S rRNA maturation in hot3 further activated RNAi to generate RDR1- and DCL2/4-dependent risiRNAs mainly from a 3’ portion of 18S rRNA. We further showed that risiRNAs in hot3 were predominantly localized in ribosome-free fractions not responsible for the 18S rRNA maturation or translation initiation defects in hot3. Our study uncovered the molecular function of HOT3/eIF5B1 in 18S rRNA maturation at the late-40S assembly stage and revealed the regulatory crosstalk among ribosome biogenesis, mRNA translation initiation, and siRNA biogenesis in plants.
2023-04-19 | GSE223171 | GEO
Project description:Iron-rich pelagic aggregates (iron snow) were collected directly onto silicate glass filters using an electronic water pump installed below the redoxcline. RNA was extracted and library preparation was done using the NEBNext Ultra II directional RNA library prep kit for Illumina. Data was demultiplied by GATC sequencing company and adaptor was trimmed by Trimgalore. After trimming, data was processed quality control by sickle and mRNA/rRNA sequences were sorted by SortmeRNA. mRNA sequences were blast against NCBI-non redundant protein database and the outputs were meganized in MEGAN to do functional analysis. rRNA sequences were further sorted against bacterial/archeal 16S rRNA, eukaryotic 18S rRNA and 10,000 rRNA sequences of bacterial 16S rRNA, eukaryotic 18S rRNA were subset to do taxonomy analysis.
2021-10-15 | E-MTAB-9686 | biostudies-arrayexpress
Project description:Purpose: Pre-ribosomal RNA is cleaved at defined sites, but many endonucleases involved in 18S rRNA release are not known. We apply an in vivo cross-linking technique coupled with deep sequencing (CRAC) that captures transcriptome-wide interactions between a yeast candidate pre-rRNA endonuclease (Utp24) and its targets in a living cell. Methods: We apply CRAC to an HTP-tagged Utp24 protein (HTP: His6 - TEV cleavage site - two copies of the z-domain of Protein A). At least two independent experiments were performed and analyzed separately. Results: We found that yeast Utp24 UV-crosslinked in vivo to the U3 snoRNA and all (pre-)rRNA elements that form the central pseudoknot in the 18S rRNA. The pseudoknot is an evolutionarily highly conserved structure that is required to ensure pre-rRNA processing at three cleavage sites (A0, A1 and A2) and still present in the mature rRNA. According to our crosslinking data, the endonuclease Utp24 is placed in close proximity to site A1 at the 5'-end of the 18S rRNA. Conclusion: Our study strongly supports the hypothesis that Utp24 cleaves pre-rRNA at sites A1 and A2. Examination of targets for pre-rRNA endonucleases in yeast cells.
2016-04-01 | E-GEOD-75991 | biostudies-arrayexpress