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Series GSE143169 Query DataSets for GSE143169
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
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.
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.
Contributor(s) Dai L, Gu S
Citation(s) 32819190
Submission date Jan 06, 2020
Last update date Apr 04, 2022
Contact name Lisheng Dai
Phone 3018466653
Organization name NCI-Frederick
Street address 1050 Boyles St.
City Frederick
State/province MD
ZIP/Postal code 21702
Country USA
Platforms (2)
GPL15520 Illumina MiSeq (Homo sapiens)
GPL21311 PacBio RS II (Homo sapiens)
Samples (20)
GSM4251590 293T_PacBio_Sequencing
GSM4251591 HeLa_PacBio_Sequencing
GSM4251592 MCF7_PacBio_Sequencing
BioProject PRJNA599131
SRA SRP239556

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Supplementary file Size Download File type/resource
GSE143169_RAW.tar 24.9 Mb (http)(custom) TAR (of FASTA, TXT)
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Raw data are available in SRA
Processed data provided as supplementary file

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