Genome-wide chromatin conformation capture (Hi-C) of primary murine nephron progenitor cells
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE222062 | GEO | 2024/03/21
REPOSITORIES: GEO
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Project description:Genome-wide chromatin conformation capture (Hi-C) of primary murine nephron progenitor cells
| PRJNA917552 | ENA
Project description:Nephron progenitor cells (NPCs) self-renew and differentiate into nephrons, the functional units of the kidney. Here we report manipulation of p38 and YAP activity creates a synthetic niche that allows the long-term clonal expansion of primary mouse and human NPCs, and induced NPCs (iNPCs) from human pluripotent stem cells. Cultured iNPCs resemble closely primary human NPCs, generating nephron organoids with abundant distal convoluted tubule cells, which are not observed in published kidney organoids. The synthetic niche reprograms differentiated nephron cells into NPC state, recapitulating the plasticity of developing nephron in vivo. Scalability and ease of genome-editing in the cultured NPCs allow for genome-wide CRISPR screening, identi-fying novel genes associated with kidney development and disease. A rapid, efficient, and scala-ble organoid model for polycystic kidney disease was derived directly from genome-edited NPCs, and validated in drug screen. These technological platforms have broad applications to kidney development, disease, plasticity, and regeneration.
2024-03-14 | GSE251862 | GEO
Project description:This series contains ATAC-Seq data generated from mouse nephron progenitor cells (NPCs) isolated from different strains/mutants and developmental timepoints as indicated.
2024-03-21 | GSE224324 | GEO
Project description:Chromosome conformation capture (3C)-based assays are used to map chromatin interactions genome-wide. Quantitative analyses of chromatin interaction maps can lead to insights into the spatial organization of chromosomes and the mechanisms by which they fold. A number of protocols such as in situ Hi-C and Micro-C are now widely used and these differ in key experimental parameters including cross-linking chemistry and chromatin fragmentation strategy. To understand how the choice of experimental protocol determines the ability to detect and quantify aspects of chromosome folding we have performed a systematic evaluation of experimental parameters of 3C-based protocols. We find that different protocols capture different 3D genome features with different efficiencies. First, the use of crosslinkers such as DSG in addition to formaldehyde improves signal-to-noise allowing detection of thousands of additional loops and strengthening compartment signal. Second, fragmenting chromatin to the level of nucleosomes using MNase allows detection of more loops. On the other hand, protocols that generate larger multi-kb fragments produce stronger compartmentalization signals. We confirmed our results in multiple cell states such as pluripotent and differentiated cells as well as cell cycle stages; Mitosis and G1. Based on these insights we developed Hi-C 3.0, a single protocol that can be used to both efficiently detect chromatin loops and to quantify compartmentalization. Finally, this study produced ultra-deeply sequenced reference interaction maps using in situ Hi-C, Micro-C and Hi-C 3.0 for commonly cell lines in the 4D Nucleome Project.
2021-03-08 | GSE165894 | GEO
Project description:Bulk ATAC_seq on GFP expressing FACS-isolated cells from E16.5 and P0 Six2TGC and compound Ezh1 and Ezh2 mutant kidneys( Six2TGC_Ezh2-/- , Ezh1+/-; Six2TGC_Ezh2-/-; and Ezh1-/- ; scRNA-seq analysis of NPCs (Six2/GFP+ cells) from E16.5 as well as P2 kidneys. Genotypes analyzed: Six2TGC (E16.5&P2), Six2TGCEzh2-/- (E16.5), Ezh1+/-;Six2TGCEzh2-/- (E16.5), and Ezh2-/-;Six2TGCEzh2-/- (E16.5) Six2/GFP+ nephron progenitor cells (NPCs) give rise to all epithelial cell types of the nephron, the filtering unit of the kidney. NPCs have a limited lifespan and are consumed near the time of birth. Pre-term birth or prenatal stress further shorten the lifespan of NPCs and result in nephron deficit and chronic kidney disease. Accordingly, there is a pressing need to better understand the factors that regulate NPC lifespan in order to develop novel regenerative strategies. Epigenetic factors are implicated in maintenance of organ-restricted progenitors such as NPCs, but the chromatin-based mechanisms are not well understood. In this study, we examined the role of the H3K27 methyltransferases, Enhancer of zeste (Ezh1 and Ezh2), in NPC maintenance using a combination of gene targeting, chromatin profiling, and single-cell RNA analysis. We find that Ezh2 expression correlates with NPC growth potential, and that Ezh2 is the dominant H3K27 methyltransferase in NPCs and epithelial descendants. Surprisingly, NPCs lacking H3K27me3 maintain their progenitor state albeit cycle slowly leading to formation of fewer nephrons. Unlike Ezh2 loss-of-function, dual inactivation of Ezh1 and Ezh2 triggers overexpression of the transcriptional repressor Hey1 and downregulation of Six2 and result in unscheduled activation of Wnt4-driven differentiation, early termination of nephrogenesis and severe renal dysgenesis. Double-mutant NPCs also overexpress the Six family member, Six1. However, in this context, Six1 is unable to access the closed Six2 enhancer and fails to maintain NPC stemness. At the chromatin level, Ezh1 and Ezh2 act by restricting accessibility of AP1 factors to their genomic binding motifs and their absence provokes a regulatory landscape akin of differentiated and non-lineage cells. We conclude that Ezh2 is required for NPC renewal potential, while maintenance of NPC lifespan and tempering the differentiation program require cooperation of both Ezh1 and Ezh2.
2020-06-22 | GSE144384 | GEO
Project description:This series contains scRNA-Seq data generated using the 10x Genomcis chromium v2 platform for embryonic day 14 and postnatal day 0 mouse kidneys isolated from three different genotypes representing control and elevated nephron number mouse phenotypes.
2021-09-24 | GSE173265 | GEO
Project description:Background: Nephron progenitor cells (NPCs) undergo a stepwise process to generate all mature nephron structures. Mesenchymal to epithelial transition (MET) is considered a multi-step process of NPC differentiation to ensure progressive establishment of new nephrons. However, despite this important role, to date, no marker for NPCs undergoing MET in the nephron exists. Results: Here, we identify LGR6 as a NPC marker, expressed in very early cap mesenchyme, pre-tubular aggregates, renal vesicles and in segments of S-shaped bodies, following the trajectory of MET. By using a lineage tracing approach in embryonic explants in combination with confocal imaging and single-cell RNA sequencing, we provide evidence for the multiple fates of LGR6+ cells during embryonic nephrogenesis. Moreover, by using long-term in vivo lineage tracing, we show that postnatal LGR6+ cells are capable of generating the multiple lineages of the nephrons. Conclusion: Given the profound early mesenchymal expression and MET signature of LGR6+ cells, together with the lineage tracing of mesenchymal LGR6+ cells, we conclude that LGR6+ cells contribute to all nephrogenic segments by undergoing MET. LGR6+ cells can therefore be considered an early committed NPC population during embryonic and postnatal nephrogenesis with potential regenerative capability.
2021-05-25 | GSE164795 | GEO
Project description:Metabolic pathways driving differentiation into nephron progenitor cells (NPCs) and renal organoids from the pluripotency stage remain poorly understood. In this study, we systematically performed comprehensive metabolite and transcriptome profiling of human pluripotent stem cells (hPSCs) during differentiation into NPCs and further into multicellular organoids. We found activation of distinct metabolic pathways during early-stage progression from hPSCs to NPCs; however, later-stage differentiation from NPCs to organoids, and the intervening developmental stages between them, largely shared similar metabolic profiles. Among the pathways changing in early differentiation, the alanine-aspartate-glutamate and glutamine pathways were found to be significantly altered by both an enrichment and by pathway impact analysis. Moreover, hPSCs survived glutamine deprivation during in vitro differentiation, and NPCs were successfully generated both in the absence and presence of glutamine and glutamate. Surprisingly, glutamine deprivation resulted in enhanced maturation, by accelerating PAX8 expression, and enriching for podocytes as detected through single cell RNA-Seq analysis. Taken together, these findings highlight a critical regulatory role of glutamine metabolism in the derivation of nephron progenitor cells and renal organoids and identify a controlling metabolic pathway of early renal differentiation.
2022-06-01 | GSE128937 | GEO
Project description:The aggregation of nephron progenitor cells (NPC) is required to form the nephron precursor renal vesicle (RV) as they undergo the mesenchymal-to-epithelial transition (MET). Canonical Wnt signaling regulates the MET of NPCs via the effector molecule β-catenin. β-catenin acts as a transcriptional co-activator during the differentiation of NPCs, however, if it has any role as an adhesion regulating molecule via the cadherin-catenin complex is currently unknown. Using in vitro NPC culture system with gene editing and modulated Wnt signaling input, we investigated the morphological transition of NPCs and its underlying mechanism modeling the initial step of the MET in vivo. Increasing Wnt signaling input with the small molecule agonist CHIR resulted in the aggregation of NPCs similar to the generation of the nephron anlagen. By removing β- and α-catenin from the NPCs, NPCs are sorted out from these aggregates, demonstrating the key role of the cadherin-catenin complex in the aggregation. β-catenin was essential for the induction, however, the removal of α-catenin did not affect the transcriptional profile of NPCs. Removal of extracellular Ca2+ resulted in the transient loss of cell-cell contacts suggesting the role of cadherins in the aggregation process. The analysis of in vitro bulk RNA-seq cadherin expression profile matched the in vivo cadherin expression profile determined by single-cell RNA-seq. The combined removal of pre-existing and inducible cadherins phenocopied the results of β- and α-catenin KO experiments highlighting the crucial role of the cadherin-catenin complex by multiple lines of evidence. Notably, the induction of NPCs is independent of their aggregation. The in vitro modeling of nephron development provides a mechanistic understanding how cell adhesion is regulated via the cadherin-catenin complex during nephrogenesis.
2024-07-29 | GSE232606 | GEO
Project description:Kidneys from multiple litters of Six2GFP+ E14.5 mouse embryos were pooled into three replicate tubes and dissociated in parallel. Six2GFP+ cells were isolated and processed for single cell sequencing using 10x Genomics technology. This resulted in a dataset of 7844 single cells representing the nephron progenitor population of the mouse kidney.
2019-05-30 | GSE130606 | GEO