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| Status |
Public on Jan 01, 2022 |
| Title |
Novel, abundant Drosha isoforms are deficient in miRNA processing in cancer cells |
| Organism |
Homo sapiens |
| Experiment type |
Non-coding RNA profiling by high throughput sequencing
Other
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| Summary |
MicroRNAs (miRNAs) are a class of small noncoding RNAs about 22-nucleotide (nt) in length that collectively regulate more than 60% of coding genes. Aberrant miRNA expression is associated with numerous diseases, including cancer. miRNA biogenesis is licensed by the ribonuclease (RNase) III enzyme Drosha, the regulation of which is critical in determining miRNA levels. We and others have previously revealed that alternative splicing regulates the subcellular localization of Drosha. To further investigate the alternative splicing landscape of Drosha transcripts, we performed PacBio sequencing in different human cell lines. We identified two novel isoforms resulting from partial intron-retention in the region encoding the Drosha catalytic domain. One isoform (AS27a) generates a truncated protein that is unstable in cells. The other (AS32a) produces a full-length Drosha with a 14 amino acid insertion in the RIIID domain. By taking advantage of Drosha knockout cells in combination with a previously established reporter assay, we demonstrated that Drosha-AS32a lacks cleavage activity. Furthermore, neither Drosha-27a nor Drosha-32a were able to rescue miRNA expression in the Drosha knockout cells. Interestingly, both isoforms were abundantly detected in a wide range of cancer cell lines (up to 15% of all Drosha isoforms). Analysis of the RNA-seq data from over 1000 breast cancer patient samples revealed that the AS32a is relatively more abundant in tumors than in normal tissue, suggesting that AS32a may play a role in cancer development.
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| Overall design |
For Pacbio sequencing, 4 human cell lines, including HEK293T, HeLa, MCF-7, U2OS were used, and in total 4 samples. For small RNA sequencing, HEK293T and HeLa cells were used, and in total 6 samples in HEK293T cells and 2 samples in HeLa cells. There are 2 replicates for small RNA sequencing. See sample description for the details of cell lines.
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| Contributor(s) |
Dai L, Gu S |
| Citation(s) |
32819190 |
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| Submission date |
Jan 06, 2020 |
| Last update date |
Apr 04, 2022 |
| Contact name |
Lisheng Dai |
| E-mail(s) |
lisheng.dai@nih.gov
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| Phone |
3018466653
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| Organization name |
NCI-Frederick
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| Street address |
1050 Boyles St.
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| City |
Frederick |
| State/province |
MD |
| ZIP/Postal code |
21702 |
| Country |
USA |
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| Platforms (2) |
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| Samples (20)
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| Relations |
| BioProject |
PRJNA599131 |
| SRA |
SRP239556 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE143169_RAW.tar |
24.9 Mb |
(http)(custom) |
TAR (of FASTA, TXT) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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