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Next Generation Sequencing Facilitates Quantitative Analysis of normal and YTHDF2 Overexpressing SMMC7721 cells in normoxia and hypoxia condition

ABSTRACT: Next Generation Sequencing Facilitates Quantitative Analysis of normal and YTHDF2 Overexpressing SMMC7721 cells in normoxia and hypoxia condition

PROVIDER: PRJNA493747 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR7939323_1.fastq.gz	Fastqsanger.gz
	SRR7939323_2.fastq.gz	Fastqsanger.gz
	SRR7939324_1.fastq.gz	Fastqsanger.gz
	SRR7939324_2.fastq.gz	Fastqsanger.gz
	SRR7939325_1.fastq.gz	Fastqsanger.gz

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Next Generation Sequencing Facilitates Quantitative Analysis of normal and YTHDF2 Overexpressing SMMC7721 cells in normoxia and hypoxia condition

Project description:Dynamic N6-methyladenosine (m6A) modification was previously identified as a ubiquitous post-transcriptional regulation that affected mRNA homeostasis. However, the m6A-related epitranscriptomic alterations and functions remain elusive in human diseases such as cancer. Here we show that hypoxia restrains the ‘reader’ protein YTH domain family 2 (YTHDF2), to stabilize the methylated oncogene mRNAs in inflammation-associated liver cancer. YTHDF2 silenced in human cancer cells or depleted in mouse hepatocytes evoked pro-inflammatory responses and accelerated tumor growth and metastatic progression. Under hypoxia, YTHDF2 processed the decay of m6A-containing interleukin 11 (IL11) and serpin family E member 2 (SERPINE2) mRNAs, which were responsible for the inflammation-mediated malignancy. Reciprocally, YTHDF2 transcription succumbed to hypoxia-inducible factor-2α (HIF-2α). Administration of a HIF-2α antagonist (PT2385) restored YTHDF2-programed epigenetic machinery and repressed liver cancer. Hence, our findings provide a new insight into the mechanism by which hypoxia adapts m6A-mRNA editing to promote cancer.

2020-03-17 | GSE120611 | GEO

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Expression profile of select glucose and lipid metabolic genes mediated by STIM1 knockout, or STIM1 knockout plus Snail1 overexpression in SMMC7721 cells

Project description:To investigate the altered glucose and lipid metabolism genes after STIM1 knockout (STIM1 KO) or STIM1 knockout plus Snail1 OE (STIM1 KO+Snail1 OE) in SMMC7721 cells. STIM1 KO-SMMC7721 cells were constructed by CRISPR/Cas9, and STIM1 KO+Snail1 OE-SMMC7721 cells was established via lentiviral Infection of Snail1 overexpression in STIM1 KO-SMMC7721 cells. The cells mentioned above were cultured in DMEM supplemented with 10% (v/v) FBS at 37 °C in 5% CO2. When the cells grow to 60~70% confluent, total RNA was extracted by using Trizol (TAKARA, Japan). Reverse transcription was performed from 1μg total RNA using PrimeScript™ RT reagent Kit with gDNA Eraser (TAKARA), Quantitative real-time PCR were performed (CFX386, BioRad) with 40 cycles at 95 °C for 10 seconds, 59 °C for 20 seconds and 72 °C for 30 seconds.

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Expression profile of select glucose and lipid metabolic genes mediated by STIM1 knockout in SMMC7721 cells

Project description:To investigate the altered glucose and lipid metabolic genes after STIM1 knockout in SMMC7721 cells. STIM1 knockout SMMC7721 cells were constructed by CRISPR/Cas9. The cells mentioned above were cultured in DMEM supplemented with 10% (v/v) FBS at 37°C in 5% CO2. When the cells grow to 60~70% confluent, Total RNA was extracted by using Trizol (TAKARA, Japan). Reverse transcription was performed from 1μg total RNA using PrimeScript™ RT reagent Kit with gDNA Eraser (TAKARA), Quantitative real-time PCR were performed (CFX386, BioRad) with 40 cycles at 95 °C for 10 seconds, 59 °C for 20seconds and 72°C for 30 seconds.

2019-08-17 | GSE135900 | GEO

Comparision of gene expression in human nucleus pulposus cells under hypoxia and normoxia condition

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Expression profile of select glucose and lipid metabolic genes mediated by STIM1-Snail1 circuit in SMMC7721 cells

Project description:To investigate the altered glucose and lipid metabolic genes after Snail1 overexpression, Snail1 plus STIM1 double overexpression in SMMC7721 cells. Snail1 overexpression (Snail1 OE)- and Snail1 plus STIM1 double overexpression(Snail1+STIM1 dOE)- SMMC7721 cells were established via lentiviral Infection. The cells mentioned above were cultured in DMEM supplemented with 10% (v/v) FBS at 37°C in 5% CO2. When the cells grow to 60~70% confluent, Total RNA was extracted by using Trizol (TAKARA, Japan). Reverse transcription was performed from 1μg total RNA using PrimeScript™ RT reagent Kit with gDNA Eraser (TAKARA), Quantitative real-time PCR were performed (CFX386, BioRad) with 40 cycles at 95 oC for 10 seconds, 59 oC for 20seconds and 72 oC for 30 seconds.

2019-08-17 | GSE135901 | GEO

SLAM-Seq analysis of Ythdf2-deficient pre-leukemic cells

Project description:The mRNA m6A reader YTHDF2 is overexpressed in a broad spectrum of human acute myeloid leukemias (AML). To study the role of YTHDF2 on mRNA decay rates in leukemia, c-Kit+ cells from foetal livers of Ythdf2fl/fl; Vav-iCre (Ythdf2CKO) and Ythdf2fl/fl (Ythdf2CTL) 14.5 dpc embryos were transduced with Meis1 and Hoxa9 oncogenes and serially re-plated to generate pre-leukemic cells. Medium with 4SU was used for pre-leukemic cells labelling for 12 hours and was later replaced with 4SU-free medium (time 0). Cells were collected immediately after medium change and at 1, 3 and 9 hours for library generation. RNA from Ythdf2CKO (n=3 biological replicates) and Ythdf2CTL (n=3 biological replicates) pre-leukemic cells were used for SLAM-seq library generation.

2019-04-05 | E-MTAB-7784 | biostudies-arrayexpress

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.

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