GSE39855
GSE GEOLIN28 binds messenger RNAs at GGAGA motifs and regulates splicing factor abundance (splicing array)
Relations
Summary
LIN28 is a conserved RNA binding protein implicated in pluripotency, reprogramming and oncogenesis. Previously shown to act primarily by blocking let-7 microRNA (miRNA) biogenesis, here we elucidate distinct roles of LIN28 regulation via its direct messenger RNA (mRNA) targets. Through cross-linking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq) in human embryonic stem cells and somatic cells expressing exogenous LIN28, we have defined discrete LIN28 binding sites in a quarter of human transcripts. These sites revealed that LIN28 binds to GGAGA sequences enriched within loop structures in mRNAs, reminiscent of its interaction with let-7 miRNA precursors. Among LIN28 mRNA targets, we found evidence for LIN28 autoregulation and also direct but differing effects on the protein abundance of splicing regulators in somatic and pluripotent stem cells. Splicing-sensitive microarrays demonstrated that exogenous LIN28 expression causes widespread downstream alternative splicing changes. These findings identify important regulatory functions of LIN28 via direct mRNA interactions.
Overall Design
In triplicate, polyA-selected RNA was extracted from untreated Flp-In-293 cells, stable LIN28V5 293 cells, TDP-43 over-expressed Flp-In-293 cells, control over-expressed Flp-In-293 cells, LIN28 depleted hES cells, and control depleted hES cells, and hybridized to custom human splicing sensitive microarrays
Analysis (3 steps)
View Data Processing- Data processed using Affymetrix package (Affy Power Tools) apt-probeset-summarize.
- Iter-plier algorithm used to quantify probesets.
- HJAY_r2.pgf