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Identify mRNAs binding with IGF2BP1 in PLC-8024 cells [RIP-seq]

ABSTRACT: To investigate mRNAs binding with IGF2BP1 in HCC, we employed the RIP-seq

ORGANISM(S): Homo sapiens

PROVIDER: GSE206504 | GEO | 2023/02/28

REPOSITORIES: GEO

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Identify mRNAs binding with IGF2BP1 in PLC-8024 cells [RIP-seq]

Project description:Identify mRNAs binding with IGF2BP1 in PLC-8024 cells [RIP-seq]

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Effect of depletion of PRMT3 and IGF2BP1 on gene expression in PLC-8024 cells [PRMT3 IGF2BP1 KD RNA-seq]

Project description:To investigate the function PRMT3 and IGF2BP1 in HCC, we established PLC-8024 cell lines in which each target gene has been knocked down by shRNA.

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Transcriptome-wide identification of IGF2BP binding RNAs by RIP-seq.

Project description:We identified the mRNA targets of the insulin-like growth factor-2 (IGF2) mRNA-binding proteins 1, 2, and 3 (IGF2BP1/2/3) by RNA immunoprecipitation and sequencing (RIP-seq). HEK293T cells transfected with Flag-tagged IGF2BP1/2/3 plasmids were expanded and UV-crosslinked before harvest. We performed RIP of individual IGF2BP using anti-Flag antibody from nuclear extractions, and identified the associated mRNAs by next generation sequencing. More than 5000 transcripts, including protein coding and non-coding transcripts, were identified from each RIP-seq sample.

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Identification of LC3B's binding mRNAs through a n-RIP-seq

Project description:To identify RNAs that bind to LC3B, we employed n-RIP-seq. After obtaining the nRIP-seq data and applying a rigorous threshold for peak calling, we detected 5,285 significant peaks associated with LC3B. The coverage analysis revealed a mean coverage of 99% for LC3B-associated peaks, suggesting widespread binding throughout the transcriptome

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Identify mRNA with m6A modification in PLC-8024 cells [MeRIP-seq]

Project description:To investigate the m6A mapping in HCC, we employed MeRIP-seq to identify mRNAs with m6A modification

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The RNA-seq of HCC specimens and the RIP-seq of HNRNPD in PLC cells and WT (DMSO or Compound C) and circLARP1B-Def (DMSO) PLC/PRF/5 cells

Project description:To investigate functional transcripts in metastatic HCC, we performed high-throughput RNA sequencing (RNA-seq) of tumors from 3 metastatic HCC and 3 non-metastatic HCC. And we performed RIP-seq human PLC/PRF/5 cells to investigate the HNRNPD binding transcripts. To investigate function of circLARP1B on AMPK pathway, we performed high-throughput RNA sequencing (RNA-seq) of WT (DMSO or Compound C) and circLARP1B-Def (DMSO) PLC/PRF/5 cells.

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m6A modification profile in primary microglia with/without Igf2bp1 knockdown

Project description:Our study demonstrated that the expression of Igf2bp1 in activated microglia was significantly up-regulated, implying a role of Igf2bp1 in LPS-induced m6A modifications in microglia. To understand the roles of Igf2bp1 on LPS-induced m6A modification in microglia, we performed Igf2bp1 loss-of-function (LOF) approach. Microglia stimulated by LPS were transfected with either scrambled siRNA control or Igf2bp1 siRNA for 48 hours. To m6A modification profiles in control and Igf2bp1 LOF microglia were determined by MeRIP-seq analysis.

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Tumor suppressive function of IGF2BP1 in gastric cancer through decreasing MYC

Project description:Gastric cancer is one of the most common causes of cancer-related death worldwide. The N6-methyladenosine (m6A) reader IGF2BP1 (insulin-like growth factor-2 mRNA binding protein 1) has been reported to promote cancer progression by stabilizing oncogenic mRNAs through its m6A-binding activity in some tumors. However, the role of IGF2BP1 in gastric carcinogenesis remains unclear. In this study, we find that IGF2BP1 is significantly downregulated in tumor tissues from patients with gastric cancer. Lower expression of IGF2BP1 is associated with poor prognosis. IGF2BP1 suppresses gastric cancer cell proliferation in an m6A-dependent manner. Additionally, IGF2BP1 is able to significantly attenuate tumor growth of gastric cancer cells. Further m6A sequencing and m6A-RIP (RNA immunoprecipitation) assays show that MYC (c-myc proto-oncogene) mRNA is a target transcript of IGF2BP1 in gastric cancer cells. IGF2BP1 inhibits gastric cancer cell proliferation by reducing the mRNA and protein expression of MYC. Mechanistically, IGF2BP1 promotes the degradation of MYC mRNA and inhibits its translation efficiency. Taken together, these data suggest that IGF2BP1 plays a tumor-suppressive role in gastric carcinogenesis by downregulating MYC in an m6A-dependent manner, thereby making the IGF2BP1-MYC axis a potential target for gastric cancer treatment.

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Anti YTHDF2 RIP-seq to identify YTHDF2 target mRNAs in P0 mouse retinas

Project description:To identify the target mRNAs of the m6A reader protein YTHDF2, we carired out anti YTHDF2 RNA Immunoprecipitation (RIP) followed by RNA-sequencencing. Using EZ-Magna RIP™ RNA-Binding Protein Immunoprecipitation Kit (Millipore), RNA from P0 wild type mouse retinas was pulled down by rabbit polyclonal anti-YTHDF2 (proteintech) and then sequenced on Illumina HiSeq3000 platform. The filtered reads were mapped to the mouse reference genome (GRCm38) using STAR v2.5 with default parameters. The resulting bam files were fed to HTSeq tool to count the number of RNA-seq reads, which was further normalized to calculate FPKM. To determine which gene is enriched, we computed the FPKM from RIP elute to input and any fold change greater than 2 was considered enriched. Finally, Biological replicates of anti-YTHDF2 RIP-Seq identified 1639 transcripts. This study provides a gene list which shows mRNA binding with YTHDF2 in mouse retina.

2022-03-15 | GSE145390 | GEO

Anti-YTHDF2 RIP-Seq to identify YTHDF2 target mRNAs in hippocampus

Project description:To identify the target mRNAs of the m6A reader protein YTHDF2 in mouse hippocampus, we carired out anti YTHDF2 RNA Immunoprecipitation (RIP) followed by RNA-sequencencing. Using EZ-Magna RIP™ RNA-Binding Protein Immunoprecipitation Kit (Millipore), RNA from P40 wild type mouse hippocampus was pulled down by rabbit polyclonal anti-YTHDF2 (proteintech) and then sequenced on Illumina Novaseq 6000. The filtered reads were aligned to the mouse reference genome (GRCm38) using BWA mem (v 0.7.12).Then the MACS2 (version 2.1.0) peak calling software was used to identify regions of IP enrichment over background, followed by the motif detected by Homer (Heinz et al., 2010). Peak related genes are then confirmed by PeakAnnotator. Different peak analysis was based on the fold enrichment of peaks of different experiments. A peak was determined as different peak when the odds ratio between two groups was more than 2. Using the same method, genes associated with different peaks were identified. Finally, Biological replicates of anti-YTHDF2 RIP-Seq identified 408 mRNAs transcripts. This study provides gene lists which shows mRNA binding with YTHDF2 in mouse hippocampus.

2023-01-19 | GSE171791 | GEO

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