CRISPR activation enables high fidelity reprogramming into human pluripotent stem cells
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA682159 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13182616.fastq.gz | Fastqsanger.gz | |||
| SRR13182617.fastq.gz | Fastqsanger.gz | |||
| SRR13182618.fastq.gz | Fastqsanger.gz | |||
| SRR13182619.fastq.gz | Fastqsanger.gz | |||
| SRR13182620.fastq.gz | Fastqsanger.gz |
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Project description:CRISPR activation enables high fidelity reprogramming into human pluripotent stem cells (single cells)
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Project description:CRISPR activation enables high fidelity reprogramming into human pluripotent stem cells (bulk cells)
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Project description:Conventional reprogramming methods rely on the ectopic expression of transcription factors to reprogram somatic cells into induced pluripotent stem cells (iPSC). Forced expression of transcription factors might lead to off-target gene activation and heterogeneous reprogramming, resulting in the emergence of alternative cell types and aberrant iPSC. Activation of endogenous pluripotency factors by CRISPR activation (CRISPRa) can potentially reduce this heterogeneity. Here we describe a high efficiency reprogramming of human somatic cells into iPSC using optimized CRISPRa. Efficient reprogramming was dependent on the additional targeting of the embryo genome activation-enriched Alu-motif and the miR-302/367 locus. Single-cell transcriptome analysis revealed that the optimized CRISPRa reprogrammed cells with more directly and specifically into the pluripotent state when compared to the conventional reprogramming method. These findings support the use of CRISPRa for high quality reprogramming of human iPSC. This SuperSeries is composed of the SubSeries listed below.
2021-12-23 | GSE162530 | GEO
Project description:Conventional reprogramming methods rely on the ectopic expression of transcription factors to reprogram somatic cells into induced pluripotent stem cells (iPSC). Forced expression of transcription factors might lead to off-target gene activation and heterogeneous reprogramming, resulting in the emergence of alternative cell types and aberrant iPSC. Activation of endogenous pluripotency factors by CRISPR activation (CRISPRa) can potentially reduce this heterogeneity. Here we describe a high efficiency reprogramming of human somatic cells into iPSC using optimized CRISPRa. Efficient reprogramming was dependent on the additional targeting of the embryo genome activation-enriched Alu-motif and the miR-302/367 locus. Single-cell transcriptome analysis revealed that the optimized CRISPRa reprogrammed cells with more directly and specifically into the pluripotent state when compared to the conventional reprogramming method. These findings support the use of CRISPRa for high quality reprogramming of human iPSC.
2021-12-23 | GSE162529 | GEO
Project description:Conventional reprogramming methods rely on the ectopic expression of transcription factors to reprogram somatic cells into induced pluripotent stem cells (iPSC). Forced expression of transcription factors might lead to off-target gene activation and heterogeneous reprogramming, resulting in the emergence of alternative cell types and aberrant iPSC. Activation of endogenous pluripotency factors by CRISPR activation (CRISPRa) can potentially reduce this heterogeneity. Here we describe a high efficiency reprogramming of human somatic cells into iPSC using optimized CRISPRa. Efficient reprogramming was dependent on the additional targeting of the embryo genome activation-enriched Alu-motif and the miR-302/367 locus. Single-cell transcriptome analysis revealed that the optimized CRISPRa reprogrammed cells with more directly and specifically into the pluripotent state when compared to the conventional reprogramming method. These findings support the use of CRISPRa for high quality reprogramming of human iPSC.
2021-12-23 | GSE162528 | GEO
Project description:Conventional reprogramming methods rely on the ectopic expression of transcription factors to reprogram somatic cells into induced pluripotent stem cells (iPSCs). The forced expression of transcription factors may lead to off-target gene activation and heterogeneous reprogramming, resulting in the emergence of alternative cell types and aberrant iPSCs. Activation of endogenous pluripotency factors by CRISPR activation (CRISPRa) can reduce this heterogeneity. Here, we describe a high-efficiency reprogramming of human somatic cells into iPSCs using optimized CRISPRa. Efficient reprogramming was dependent on the additional targeting of the embryo genome activation-enriched Alu-motif and the miR-302/367 locus. Single-cell transcriptome analysis revealed that the optimized CRISPRa reprogrammed cells more directly and specifically into the pluripotent state when compared to the conventional reprogramming method. These findings support the use of CRISPRa for high-quality pluripotent reprogramming of human cells.
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Project description:Defining how human pluripotent cell identity is controlled and in particular how naïve pluripotency is acquired during cell reprogramming is crucial for the future applications of pluripotent stem cells. However, the regulatory pathways of naïve cell reprogramming remain incompletely understood. Here, we used genome-wide CRISPR-Cas9 screening to identify novel regulators of primed to naïve pluripotent stem cell reprogramming, including genes that are essential for reprogramming and genes that normally impede reprogramming and whose targeted deletion led to enhanced reprogramming. Integrated analysis defined specific chromatin complexes and signalling pathways as critical regulators of naïve reprogramming, and were largely distinct from regulators of somatic cell reprogramming. Mechanistically, PRC1.3 and PRDM14 are jointly required to transcriptionally repress developmental and gene regulatory factors to ensure naïve cell reprogramming. Additionally, small molecule inhibitors of reprogramming impediments increased the efficiency of naïve cell reprogramming, and are of practical benefit that can improve on current reprogramming methods. Taken together, we have identified novel regulators controlling the establishment of naïve pluripotency in human cells, which will open up new ways to exploit the full potential of pluripotent stem cells. These results also provide new insights into mechanisms that destabilise and reconfigure cell identity during cell state transitions.
2022-04-06 | PXD023547 | Pride
Project description:Defining how human pluripotent cell identity is controlled and in particular how naïve pluripotency is acquired during cell reprogramming is crucial for the future applications of pluripotent stem cells. However, the regulatory pathways of naïve cell reprogramming remain incompletely understood. Here, we used genome-wide CRISPR-Cas9 screening to identify novel regulators of primed to naïve pluripotent stem cell reprogramming, including genes that are essential for reprogramming and genes that impede reprogramming and whose targeted deletion led to enhanced reprogramming. Integrated analysis defined specific chromatin complexes and signalling pathways as critical regulators of naïve reprogramming, and were largely distinct from regulators of somatic cell reprogramming. Mechanistically, PRC1.3 and PRDM14 are jointly required to transcriptionally repress developmental and gene regulatory factors to ensure naïve cell reprogramming. Additionally, small molecule inhibitors of reprogramming impediments increased the efficiency of naïve cell reprogramming, and are of practical benefit that can improve on current reprogramming methods. Taken together, we have identified novel regulators controlling the establishment of naïve pluripotency in human cells, which will open up new ways to exploit the full potential of pluripotent stem cells. These results also provide new insights into mechanisms that destabilise and reconfigure cell identity during cell state transitions.
2022-03-25 | GSE164786 | GEO
Project description:To investigate the genes and pathways involved in X-chromosome reactivation and pluripotency acquisition, we generated a whole-genome CRISPR knockout screen during female NPC reprogramming into iPSCs. We analyzed the gRNA abundance in NPCs and three different reprogramming populations: non-pluripotent, early pluripotent and late pluripotent. Pair-wise comparisons of gRNA frequencies in different populations were done to identify genes and pathways involved in cell survival in reprogramming conditions, colony formation, early pluripotency acquisition and late pluripotency/X-chromosome reactivation.
2024-08-07 | GSE236244 | GEO
Project description:We recently demonstrated the chemical reprogramming of human somatic cells to pluripotent stem cells, which provides a fundamentally new approach for cell fate manipulation. However, the utility of this chemical approach is currently hampered by the slow kinetics. Here, by screening for new small-molecule boosters and systematically optimizing the original chemical reprogramming condition, we have established a robust, chemically defined protocol. With this new protocol, the entire chemical reprogramming process is remarkably accelerated, reducing the time needed from ~50 days to a minimum of 16 days. Moreover, this new protocol enables highly reproducible and efficient generation of human pluripotent stem cells from all 17 tested donors with a reprogramming efficiency of up to 31%. Our method provides a unique and powerful strategy for human cell fate manipulation and pluripotent stem cell manufacturing for cell-based therapeutic applications.
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