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Mus musculus

ABSTRACT: Genome-wide localization of NF-E2 p45 in mouse primary megakaryocytes.

PROVIDER: PRJNA179441 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR616693.fastq.gz	Fastqsanger.gz
	SRR616694.fastq.gz	Fastqsanger.gz
	SRR616695.fastq.gz	Fastqsanger.gz
	SRR616696.fastq.gz	Fastqsanger.gz
	SRR616697.fastq.gz	Fastqsanger.gz

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Project description:Comparison of gene expression profiles of p45-/- primary megakaryocytes and WT primary megakaryocytes

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Project description:ChIP-seq assay was performed with anti-p45 antibody using in vitro cultured primary megakaryocytes to identify direct p45 target genes in megakaryocytes. This analysis revealed genes that regulate platelet function, which were not known previously to be p45-regulated. ChIP was performed using a day-3 primary culture of megakaryocytes from E13.5 fetal livers of wild-type mice with an anti-p45 antibody.

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Project description:Whole livers were collected from mouse fetuses at embryonic day 13.5 (E13.5), and single-cell suspensions were prepared by successive passage through 25-gauge needles. Fetal liver cells were maintained in RPMI1640 (Wako, Osaka, Japan) supplemented with 20% charcoal-stripped fetal bovine serum (FBS), 100 U/ml penicillin, 100µg/ml streptomycin, and 50ng/ml recombinant human thrombopoietin (TPO) (generously provided by Kyowa Hakko Kirin Co. Ltd.). Megakaryocytes were harvested for RNA purification from a day-3 culture. CD41+ cells were enriched using biotinylated anti-CD41 antibody (Serotec; clone MWReg30) and streptavidin-coupled Dynabeads (Dynal Biotech ASA) from a culture of E13.5 fetal liver cells of WT and p45–/– mice. Total RNA was purified from the sorted CD41+ cells. Gene expression was measured in primary megakaryocytes cultured from p45-/- fetal livers and wild type fetal livers at E13.5. Three independent experiments were performed.

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Project description:Introgressed variants from other species can be an important source of genetic variation because they may arise rapidly, can include multiple mutations on a single haplotype, and have often been pretested by selection in the species of origin. Although introgressed alleles are generally deleterious, several studies have reported introgression as the source of adaptive alleles-including the rodenticide-resistant variant of Vkorc1 that introgressed from Mus spretus into European populations of Mus musculus domesticus. Here, we conducted bidirectional genome scans to characterize introgressed regions into one wild population of M. spretus from Spain and three wild populations of M. m. domesticus from France, Germany, and Iran. Despite the fact that these species show considerable intrinsic postzygotic reproductive isolation, introgression was observed in all individuals, including in the M. musculus reference genome (GRCm38). Mus spretus individuals had a greater proportion of introgression compared with M. m. domesticus, and within M. m. domesticus, the proportion of introgression decreased with geographic distance from the area of sympatry. Introgression was observed on all autosomes for both species, but not on the X-chromosome in M. m. domesticus, consistent with known X-linked hybrid sterility and inviability genes that have been mapped to the M. spretus X-chromosome. Tract lengths were generally short with a few outliers of up to 2.7 Mb. Interestingly, the longest introgressed tracts were in olfactory receptor regions, and introgressed tracts were significantly enriched for olfactory receptor genes in both species, suggesting that introgression may be a source of functional novelty even between species with high barriers to gene flow.

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Project description:To identify p45 target genes, we conducted gene expression profiling with p45-null megakaryocytes cultured from E14.5 fetal liver. Many genes encoding membrane proteins and enzymes related to platelet function, including Txas, Glycoprotein 6 (Gp6) and Selectin P (Selp), were repressed in the absence of p45. Considering the similar DNA binding specificity of p45 and Nrf2 in vitro, we expected p45 to activate cytoprotective genes that are established Nrf2 targets. However, the expression of numerous detoxifying enzymes and stress-responsive genes, including NAD(P)H:quinone oxidoreductase (Nqo1), were increased in the absence of p45.

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