Mountain View
biomedical and healthCAre Data Discovery Index Ecosystem
help Advanced Search
Title: Antisense Transcripts Impede Uridylation-induced Exonucleolytic Degradation to Generate Guide RNAs      
dateReleased:
02-08-2016
description:
Small non-coding RNA biogenesis typically involves cleavage of structured precursors by RNase III-like endonucleases. However, guide RNAs that direct U-insertion/deletion mRNA editing in mitochondria of trypanosomes maintain 5′ triphosphate characteristic of transcription start site and possess U-tail indicative of 3′ processing and uridylation. Here, we identified a protein complex composed of RET1 TUTase and 3′-5′ DSS1 exonuclease, and three additional subunits. This complex, termed mitochondrial 3′ processome (MPsome), is responsible for primary uridylation of ~800-nt gRNA precursors, their processive degradation to a mature length of 50-60 nt, and secondary U-tail addition. Both strands of gRNA gene are transcribed giving rise to sense and antisense precursors of similar size. Head-to-head hybridization of these transcripts blocks symmetrical 3′-5′ degradation at the fixed distance from the double-stranded region. Together, our findings suggest a model in which gRNA is derived from the 5′ extremity of a primary molecule by uridylation-induced, antisense transcription-controlled exonucleolytic degradation. 1. we first sequenced guide RNA precursor (Gel-fractioned total cellular RNA 600-1500nt was used) transcripts from three replicates to study the their tail features and also validate observation of sense/antisense accumulation upon perturbation of pre-processing complex based on few cases. 2. we then sequenced mitochondrial small RNA and built a reference for small RNAs using our custom algorithm. We then took the mitochondrial small RNA data and uncovered the sense/antisense pair. 3. We then used CLIP-Seq data to investigate in vivo binding sites and, together with RNA IP-Seq data to understand what determine the relative abundance of sense and antisense pair of the duplex. 4. We used sequenced small mitochondrial RNA in different RNAi experiments (For RNAi experiments, 30-70nt RNA fraction was gel-isolated from total cellular RNAs) to understand which protein will affect the Utail length in mature small mitochondrial RNA.
privacy:
not applicable
aggregation:
instance of dataset
ID:
E-GEOD-71292
refinement:
raw
alternateIdentifiers:
71292
keywords:
functional genomics
dateModified:
02-22-2016
availability:
available
types:
gene expression
name:
Trypanosoma brucei
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-71292/E-GEOD-71292.raw.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-71292/E-GEOD-71292.processed.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE71292
storedIn:
Gene Expression Omnibus
qualifier:
not compressed
format:
HTML
accessType:
landing page
primary:
true
authentication:
none
authorization:
none
abbreviation:
EBI
homePage: http://www.ebi.ac.uk/
ID:
SCR:004727
name:
European Bioinformatics Institute
homePage: https://www.ebi.ac.uk/arrayexpress/
ID:
SCR:002964
name:
ArrayExpress

Feedback?

If you are having problems using our tools, or if you would just like to send us some feedback, please post your questions on GitHub.