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Project description:The retina, the accessible part of the central nervous system, has served as a model system to study the relationship between energy utilization and metabolite supply. When the metabolite supply cannot match the energy demand, retinal neurons are at risk of death. As the powerhouse of eukaryotic cells, mitochondria play a pivotal role in generating ATP, produce precursors for macromolecules, maintain the redox homeostasis, and function as waste management centers for various types of metabolic intermediates. Mitochondrial dysfunction has been implicated in the pathologies of a number of degenerative retinal diseases. It is well known that photoreceptors are particularly vulnerable to mutations affecting mitochondrial function due to their high energy demand and susceptibility to oxidative stress. However, it is unclear how defective mitochondria affect other retinal neurons. Nuclear respiratory factor 1 (Nrf1) is the major transcriptional regulator of mitochondrial biogenesis, and loss of Nrf1 leads to defective mitochondria biogenesis and eventually cell death. Here, we investigated how different retinal neurons respond to the loss of Nrf1. We provide in vivo evidence that the disruption of Nrf1-mediated mitochondrial biogenesis results in a slow, progressive degeneration of all retinal cell types examined, although they present different sensitivity to the deletion of Nrf1, which implicates differential energy demand and utilization, as well as tolerance to mitochondria defects in different neuronal cells. Furthermore, transcriptome analysis on rod-specific Nrf1 deletion uncovered a previously unknown role of Nrf1 in maintaining genome stability.
2022-12-17 | GSE150258 | GEO
Project description:Purpose: the goal of this experiment was to identify differentially expressed genes in Six3 null, Six6 null and Six3;Six6 compound null retinas by RNAsequencing. Method: Retinas were dissected out from the following E13.5 mouse embryos: 1) WT (Six3F/F; Six6+/+); 2) Six3 KO (Six3F/F; CAGGCre-ERTM; Six6+/+); 3) Six6 KO (Six3F/F; Six6–/–); 4) DKO (Six3F/F; CAGGCre-ERTM; Six6–/–). RNA was then extracted from the retinas and profiled using RNAsequencing. Results: RNA isolated from three pairs of retinas for each genotype group (181.2-792 ng, RIN>9) was used for library preparation using KAPA RNA HyperPrep Kit with RiboErase. Sequencing was run on Illumina HiSeq 2500 in 100-bp single-end high-output mode in the Einstein Epigenomics Core Facility. About 30 million reads were generated for each sample. Each genotype group initially had three biological replicates, but one Six6 KO replicate was later removed due to over duplication. After trimming adapters with Trim Galore (v. 0.3.7), RNA-Seq reads were aligned back to mouse genome mm10 using Tophat (v. 2.0.13). The number of reads mapped back to each gene was calculated with HTseq (v.0.6.1) using Refseq gene annotation (downloaded from the UCSC genome browser in 03/17). The Cuffdiff in Cufflinks package (v. 2.2.1) was used to generate FPKM values. We identified 13498 transcripts with FPKM value >1 in at least one of samples. Deseq2 was used to determine the differentially expressed genes (DEGs) with FDR less than 0.05 as a cutoff.
2018-11-14 | GSE115258 | GEO
Project description:BACKGROUND:Mitochondrial dysfunction has been implicated in the pathologies of a number of retinal degenerative diseases in both the outer and inner retina. In the outer retina, photoreceptors are particularly vulnerable to mutations affecting mitochondrial function due to their high energy demand and sensitivity to oxidative stress. However, it is unclear how defective mitochondrial biogenesis affects neural development and contributes to neural degeneration. In this report, we investigated the in vivo function of nuclear respiratory factor 1 (Nrf1), a major transcriptional regulator of mitochondrial biogenesis in both proliferating retinal progenitor cells (RPCs) and postmitotic rod photoreceptor cells (PRs). METHODS:We used mouse genetic techniques to generate RPC-specific and rod PR-specific Nrf1 conditional knockout mouse models. We then applied a comprehensive set of tools, including histopathological and molecular analyses, RNA-seq, and electroretinography on these mouse lines to study Nrf1-regulated genes and Nrf1's roles in both developing retinas and differentiated rod PRs. For all comparisons between genotypes, a two-tailed two-sample student's t-test was used. Results were considered significant when P <?0.05. RESULTS:We uncovered essential roles of Nrf1 in cell proliferation in RPCs, cell migration and survival of newly specified retinal ganglion cells (RGCs), neurite outgrowth in retinal explants, reconfiguration of metabolic pathways in RPCs, and mitochondrial morphology, position, and function in rod PRs. CONCLUSIONS:Our findings provide in vivo evidence that Nrf1 and Nrf1-mediated pathways have context-dependent and cell-state-specific functions during neural development, and disruption of Nrf1-mediated mitochondrial biogenesis in rod PRs results in impaired mitochondria and a slow, progressive degeneration of rod PRs. These results offer new insights into the roles of Nrf1 in retinal development and neuronal homeostasis and the differential sensitivities of diverse neuronal tissues and cell types of dysfunctional mitochondria. Moreover, the conditional Nrf1 allele we have generated provides the opportunity to develop novel mouse models to understand how defective mitochondrial biogenesis contributes to the pathologies and disease progression of several neurodegenerative diseases, including glaucoma, age-related macular degeneration, Parkinson's diseases, and Huntington's disease.
| S-EPMC6192121 | biostudies-literature
Project description:This study will reveal the importance of nrf1 in revealing the translational control of Fe homeostasis
2019-12-30 | E-MTAB-8158 | biostudies-arrayexpress
Project description:Eukaryotic transcription factors (TFs) are key determinants of gene activity, yet they bind only a fraction of their corresponding DNA sequence motifs in any given cell type. Chromatin has the potential to restrict accessibility of binding sites; however, in which context chromatin states are instructive for TF binding remains mainly unknown. To explore the contribution of DNA methylation to constrained TF binding, we mapped DNase-I-hypersensitive sites in murine stem cells in the presence and absence of DNA methylation. Methylation-restricted sites are enriched for TF motifs containing CpGs, especially for those of NRF1. In fact, the TF NRF1 occupies several thousand additional sites in the unmethylated genome, resulting in increased transcription. Restoring de novo methyltransferase activity initiates remethylation at these sites and outcompetes NRF1 binding. This suggests that binding of DNA-methylationsensitive TFs relies on additional determinants to induce local hypomethylation. In support of this model, removal of neighbouring motifs in cis or of a TF in trans causes local hypermethylation and subsequent loss of NRF1 binding. This competition between DNA methylation and TFs in vivo reveals a case of cooperativity between TFs that acts indirectly via DNA methylation. Methylation removal by methylation-insensitive factors enables occupancy of methylation-sensitive factors, a principle that rationalizes hypomethylation of regulatory regions. DNase-seq (2 replicates) in mouse embryonic stem cells with (WT) and without DNA methylation (DNMT TKO). RNA-seq (3 replicates) in WT and DNMT TKO cells and in DNMT TKO cells after treatment with control siRNA or siRNA targeting Nrf1. H3K27ac ChIP-seq (2 replicates) in WT and DNMT TKO cells. NRF1 ChIP-seq (2 replicates) in WT and DNMT TKO cells, in WT upon culture in different conditions (adaptation to 2i and back to serum), upon transient overexpression of NRF1 and after differentiation into neuronal progenitor cells (NP). Whole-genome bisulfite sequencing in DNMT TKO cells and in WT upon culture in different conditions (adaptation to 2i and back to serum). NRF1 ChIP-seq (2 replicates) in human HMEC and HCC1954 cells.
2015-12-16 | E-GEOD-67867 | biostudies-arrayexpress
Project description:Purpose: Next-generation sequencing (NGS) for systematic analysis to discover Nrf1-dependent cellular pathways involved in defending liver against the stress of cholesterol excess. Methods: Liver mRNA profiles of 9-12-old wild-type (WT in fastq files; flox in paper) and liver Nrf1 deficient (KO in fastq files; L-KO in paper) mice were generated by deep sequencing, in triplicate or quadruplicate, using Illumina HiSeq2500. The sequence reads that passed quality filters were analyzed at the transcript level with TopHat followed by Cufflinks. qRT–PCR validation of selected target genes was performed using SYBR Green assays. Results: We mapped sequence reads to the mouse genome (build mm10) and identified greater than 10,000 transcripts in the liver of WT and KO mice. RNA-seq data revealed a role for Nrf1 in regulating inflammation, lipid metabolism, and sterol metabolism in response to a cholesterol challenge. Conclusion: Our study reveal Nrf1 as a guardian of cholesterol homeostasis and a core component of adaptive responses to excesses in cellular cholesterol.
2017-11-30 | GSE103949 | GEO
Project description:To investigate the role of NRF1 in regulating primordial germ cell development, We established conditional knockout mice of Nrf1 in primordial germ cell to observe the effect of Nrf1 knockout on the development of primordial germ cell. At the same time, we utilized a pluripotent stem cell differentiation system in vitro to obtain PGCL cells for chip_ Seq, analyze which genes Nrf1 directly binds to. Meanwhile, we established a pluripotent stem cell line induced by Nrf1 overexpression and performed RNA_seq analysis on PGCL cells overexpressing Nrf1 obtained in vitro
2023-08-05 | GSE231662 | GEO
Project description:To investigate the role of NRF1 in regulating primordial germ cell development, We established conditional knockout mice of Nrf1 in primordial germ cell to observe the effect of Nrf1 knockout on the development of primordial germ cell. At the same time, we utilized a pluripotent stem cell differentiation system in vitro to obtain PGCL cells for chip_ Seq, analyze which genes Nrf1 directly binds to. Meanwhile, we established a pluripotent stem cell line induced by Nrf1 overexpression and performed RNA_seq analysis on PGCL cells overexpressing Nrf1 obtained in vitro
2023-08-05 | GSE231661 | GEO
Project description:Regulation of retinal differentiation in mammals is not adequately understood. Using single-cell RNA sequencing of control and Six3 and Six6 compound-mutant mouse embryonic eye-cups, we identified cell clusters and developmental trajectories jointly regulated by transcription factor Six3 and its close paralog Six6. In control retinas, naïve retinal progenitor cells had two major trajectories leading to ciliary margin cells and retinal neurons, respectively. The ciliary margin trajectory was directly from naïve retinal progenitor cells at G1 phase whereas the neuronal trajectory was through a neurogenic state marked by Atoh7 expression. Upon Six3 and Six6 dual deficiencies, both naïve and neurogenic retinal progenitors were defective, ciliary margin differentiation was enhanced, and multi-lineage neuronal differentiation was disrupted. An ectopic neuronal trajectory lacking the Atoh7+ state led to ectopic neurons. Additionally, opposing gradients of Wnt and Fgf signaling were perturbed. Our findings provide deeper insight into molecular mechanisms underlying early retinal differentiation in mammals.
2024-08-03 | GSE241946 | GEO
Project description:Charaterize NRF1 cistrome in mouse muscle
2024-03-22 | GSE252431 | GEO