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SRP109284
Homo sapiens
6 Downloadable Samples
Illumina HiSeq 2500
Description
Immunodeficient mouse models have been valuable for studies of human hematopoiesis, but high-fidelity recapitulation of erythropoiesis in most xenograft recipients remains elusive. Recently developed immunodeficient and Kit mutant mice, however, have provided a suitable background to achieve higher-level human erythropoiesis after long-term hematopoietic engraftment. While there has been some characterization of human erythropoiesis in these models, a comprehensive analysis of various developmental stages has not yet been reported. Here, we have utilized cell surface phenotypes, morphologic analyses, and molecular studies to fully characterize human erythropoiesis from multiple developmental stages in immunodeficient and Kit mutant mouse models following long-term hematopoietic stem and progenitor cell engraftment. We show that human erythropoiesis in such models demonstrates complete maturation and enucleation, as well as developmentally appropriate globin gene expression. These results provide a framework for future studies to utilize this model system for interrogating disorders affecting human erythropoiesis and for developing improved therapeutic approaches. Overall design: (mRNA-seq) RNA-seq of human CD235a+ cells isolated 14-16 weeks post-implantation from mouse bone marrow were performed for three biological replicates each of mice xenograted with adult bone marrow-derived human CD34+ cells and cord blood-derived CD34+ cells.
Publication Title
Developmentally-faithful and effective human erythropoiesis in immunodeficient and Kit mutant mice.
Sample Metadata Fields
Specimen part, Subject
View Samples- Escherichia coli, Homo sapiens
- 6 Downloadable Samples
Affymetrix Human Gene 2.0 ST Array (hugene20st)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
Systematic Functional Dissection of Common Genetic Variation Affecting Red Blood Cell Traits.
Sample Metadata Fields
Specimen part, Cell line
View Samples- Homo sapiens
- 28 Downloadable Samples
- NextSeq 500
Description
In vitro cultured CD34+ derived erythroblasts were sorted using surface markers and processed using RNA-seq Overall design: Biological replicates (3 or 4 per population) were processed across 2-3 biological donors for 8 sorted populations for RNA-seq
Publication Title
Transcriptional States and Chromatin Accessibility Underlying Human Erythropoiesis.
Sample Metadata Fields
Subject
View Samples- Homo sapiens
- 7 Downloadable Samples
- NextSeq 500
Description
HUDEP-2 cells were lentivirally infected with CRISPRi constructs using a nontargeting guide or guides targeting an enhancer in the TMCC2 locus Overall design: Whole transcriptome libraries were sequenced for three replicates of non-targeting gRNA and two replicates each for two different gRNA targeting a regulatory region upstream of the TMCC2 erythroid-specific isoform
Publication Title
Transcriptional States and Chromatin Accessibility Underlying Human Erythropoiesis.
Sample Metadata Fields
Cell line, Subject
View Samples- Homo sapiens
- 16 Downloadable Samples
- Illumina HiSeq 2500
Description
Thousands of long non-coding RNAs (lncRNAs) have been identified in the human genome, but specific biological functions and biochemical mechanisms have been discovered for only about a dozen lncRNAs. One specific lncRNA, Non-coding RNA Activated by DNA Damage (NORAD), has recently been shown by genetic deletion to be required for maintaining genomic stability, but its molecular mechanism is unknown. Here, we combine RNA antisense purification (RAP) and quantitative mass spectrometry to identify proteins that directly interact with NORAD in living cells. We show that NORAD interacts with proteins involved in DNA replication and repair in steady-state cells and localizes to the nucleus upon stimulation with replication stress or DNA damage. In particular, NORAD interacts with RBMX (an emerging component of the DNA-damage response) and encodes the strongest RBMX-binding site in the transcriptome. We demonstrate that NORAD controls the ability of RBMX to assemble a ribonucleoprotein complex, which we term NORAD-Activated Ribonucleoprotein Complex 1 (NARC1), containing known suppressors of genomic instability: topoisomerase I (TOP1), ALYREF and the PRPF19/CDC5L complex. Cells depleted of NORAD or RBMX display an increased frequency of chromosome segregation errors, reduced replication-fork velocity and altered cell cycle progression phenotypes that are mechanistically linked to TOP1 and PRPF19/CDC5L function. Expression of NORAD in trans can rescue defects caused by NORAD depletion, but rescue is significantly impaired when the RBMX-binding site in NORAD is deleted. Our results demonstrate that the interaction between NORAD and RBMX is important for NORAD function and that NORAD is required for the assembly of a previously unknown topoisomerase complex (NARC1) that contributes to maintaining genomic stability. Moreover, we uncover a novel function for lncRNAs in modulating the ability of an RNA-binding protein to assemble a higher-order ribonucleoprotein complex. Overall design: We examined gene expression changes and alternative splicing events in wildtype and NORAD depleted cells using RNA sequencing.
Publication Title
The NORAD lncRNA assembles a topoisomerase complex critical for genome stability.
Sample Metadata Fields
Cell line, Subject, Time
View Samples- Homo sapiens
- 9 Downloadable Samples
- NextSeq 500
Description
Follow-up work was performed for SF3A2, a gene among the hits identified in a red blood cell trait GWAS-informed shRNA screen. Differential splicing effects were assayed to investigate resulting effects on the differentiating erythroid cell spliceome and explore potential modifier relationships with other known splicing defects associated with human disease. Overall design: Examination of differential splicing events resulting from knockdown of splicing factor 3a subunit 2 (SF3A2) in three unique donor CD34+ cells populations undergoing erythroid differentiation. Two shRNA targeting SF3A2 were tested, along with a negative control shRNA targeting luciferase (which should not be expressed) using paired-end sequencing.
Publication Title
Gene-centric functional dissection of human genetic variation uncovers regulators of hematopoiesis.
Sample Metadata Fields
Specimen part, Subject
View Samples- Homo sapiens
- 172 Downloadable Samples
Description
Lineage tracing provides unprecedented insights into the fate of individual cells and their progeny in complex organisms. While effective genetic approaches have been developed in vitro and in animal models, these cannot be used to interrogate human physiology in vivo. Instead, naturally occurring somatic mutations have been utilized to infer clonality and lineal relationships between cells in human tissues, but current approaches are limited by high error rates and scale, and provide little information about the state or function of the cells. Here, we show how somatic mutations in mitochondrial DNA (mtDNA) can be tracked by current single cell RNA-Seq (scRNA-Seq) or single cell ATAC-Seq (scATAC-Seq) for simultaneous analysis of single cell lineage and state. We leverage somatic mtDNA mutations as natural genetic barcodes and demonstrate their use as clonal markers to infer lineal relationships. We trace the lineage of human cells by somatic mtDNA mutations in a native context both in vitro and in vivo, and relate it to expression profiles and chromatin accessibility. Our approach should allow lineage tracing at a 100- to 1,000-fold greater scale than with single cell whole genome sequencing, while providing information on cell state, opening the way to chart detailed cell lineage and fate maps in human health and disease. Overall design: A population of 25 transfected TF1 cells were expanded and forwarded to a combination of 1) ATAC-seq and single cell RNA-seq. The single-cell RNA-seq data are listed here. Meta data includes heteroplasmic variant information per cell as well as the group assigned based on the lentiviral barcoding
Publication Title
Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 135 Downloadable Samples
Description
Lineage tracing provides unprecedented insights into the fate of individual cells and their progeny in complex organisms. While effective genetic approaches have been developed in vitro and in animal models, these cannot be used to interrogate human physiology in vivo. Instead, naturally occurring somatic mutations have been utilized to infer clonality and lineal relationships between cells in human tissues, but current approaches are limited by high error rates and scale, and provide little information about the state or function of the cells. Here, we show how somatic mutations in mitochondrial DNA (mtDNA) can be tracked by current single cell RNA-Seq (scRNA-Seq) or single cell ATAC-Seq (scATAC-Seq) for simultaneous analysis of single cell lineage and state. We leverage somatic mtDNA mutations as natural genetic barcodes and demonstrate their use as clonal markers to infer lineal relationships. We trace the lineage of human cells by somatic mtDNA mutations in a native context both in vitro and in vivo, and relate it to expression profiles and chromatin accessibility. Our approach should allow lineage tracing at a 100- to 1,000-fold greater scale than with single cell whole genome sequencing, while providing information on cell state, opening the way to chart detailed cell lineage and fate maps in human health and disease. A variety of experimental designs using cells derived from both in vitro and in vivo to determine the efficacy of using mtDNA mutations in human clonal tracing. Overall design: A population of 30 primary hematopoietic cells were expanded and forwarded to a combination of ATAC-seq and single cell RNA-seq. single cell RNA-seq samples are listed here.
Publication Title
Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 81 Downloadable Samples
- NextSeq 500
Description
Lineage tracing provides unprecedented insights into the fate of individual cells and their progeny in complex organisms. While effective genetic approaches have been developed in vitro and in animal models, these cannot be used to interrogate human physiology in vivo. Instead, naturally occurring somatic mutations have been utilized to infer clonality and lineal relationships between cells in human tissues, but current approaches are limited by high error rates and scale, and provide little information about the state or function of the cells. Here, we show how somatic mutations in mitochondrial DNA (mtDNA) can be tracked by current single cell RNA-Seq (scRNA-Seq) or single cell ATAC-Seq (scATAC-Seq) for simultaneous analysis of single cell lineage and state. We leverage somatic mtDNA mutations as natural genetic barcodes and demonstrate their use as clonal markers to infer lineal relationships. We trace the lineage of human cells by somatic mtDNA mutations in a native context both in vitro and in vivo, and relate it to expression profiles and chromatin accessibility. Our approach should allow lineage tracing at a 100- to 1,000-fold greater scale than with single cell whole genome sequencing, while providing information on cell state, opening the way to chart detailed cell lineage and fate maps in human health and disease. A variety of experimental designs using cells derived from both in vitro and in vivo to determine the efficacy of using mtDNA mutations in human clonal tracing. Overall design: Individually sorted cells from clonally derived TF1 clones (C9, D6, and G10) were processed with single cell RNA-seq (Smart-seq2)
Publication Title
Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Homo sapiens
- 3 Downloadable Samples
- NextSeq 500
Description
Lineage tracing provides unprecedented insights into the fate of individual cells and their progeny in complex organisms. While effective genetic approaches have been developed in vitro and in animal models, these cannot be used to interrogate human physiology in vivo. Instead, naturally occurring somatic mutations have been utilized to infer clonality and lineal relationships between cells in human tissues, but current approaches are limited by high error rates and scale, and provide little information about the state or function of the cells. Here, we show how somatic mutations in mitochondrial DNA (mtDNA) can be tracked by current single cell RNA-Seq (scRNA-Seq) or single cell ATAC-Seq (scATAC-Seq) for simultaneous analysis of single cell lineage and state. We leverage somatic mtDNA mutations as natural genetic barcodes and demonstrate their use as clonal markers to infer lineal relationships. We trace the lineage of human cells by somatic mtDNA mutations in a native context both in vitro and in vivo, and relate it to expression profiles and chromatin accessibility. Our approach should allow lineage tracing at a 100- to 1,000-fold greater scale than with single cell whole genome sequencing, while providing information on cell state, opening the way to chart detailed cell lineage and fate maps in human health and disease. A variety of experimental designs using cells derived from both in vitro and in vivo to determine the efficacy of using mtDNA mutations in human clonal tracing. Overall design: Cells from 3 separate TF1 clones (C9, D6, and G10) were processed with bulk RNA-seq (Smart-seq2)
Publication Title
Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples