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| Status |
Public on May 07, 2014 |
| Title |
Small RNA deep sequencing of cellular subcompartments |
| Organism |
Homo sapiens |
| Experiment type |
Non-coding RNA profiling by high throughput sequencing
|
| Summary |
RNA deep sequencing efforts have revealed abundant expression of small RNAs derived from small nucleolar (sno) RNAs. We employ here spatial RNA deep sequencing to assess the expression of 10-40 nt small RNAs in subcellular compartments of HeLa cells. Total cellular, cytoplasmic, nuclear and nucleolar fractions were isolated, RNA was purified and size-fractionated in a two-step process to yield 10-40 nt RNA fractions. cDNA libraries were constructed and sequenced on a Ion Torrent platform. Vast majority of cellular, cytoplasmic and nuclear small RNA reads were identified as miRNAs. We found the expression of eleven ten miRNAs in the nucleolar preparations using a cut-off rate of 10 reads. Several miRNAs had a greater relative abundance in the nucleolus compared to the other compartments. The nucleolar small RNAs had a unique size distribution consisting of 19-20 and 25 nt RNAs, which were predominantly composed of small snoRNA-derived box C/D RNAs (termed as sdRNA). Sequences from 47 sdRNAs were identified, which mapped both 5’ and 3’ ends of the snoRNAs, and retained conserved box C or D motifs. SdRNA reads from SNORD44 comprised 74% of all nucleolar sdRNAs, and were confirmed by Northern blotting as 20 and 25 nt RNAs. This study reveals a rich representation of cell-compartment specific expression of small RNAs and the distinctive unique composition of the nucleolar small RNAs.
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| |
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| Overall design |
Examination of small RNAs in cellular subcompartments
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| |
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| Contributor(s) |
Bai B, Laiho M, Wheelan S, Yegnasubramanian S |
| Citation(s) |
24918059 |
| Submission date |
Aug 21, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Marikki Laiho |
| Organization name |
The Johns Hopkins University School of Medicine
|
| Department |
Department of Radiation Oncology and Molecular Radiation Sciences
|
| Street address |
1550 Orleans Street, CRB II, Room 444
|
| City |
Baltimore |
| State/province |
MD |
| ZIP/Postal code |
21231 |
| Country |
USA |
| |
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| Platforms (1) |
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| Samples (4)
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| Relations |
| BioProject |
PRJNA215894 |
| SRA |
SRP028959 |
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