A novel human pluripotent stem cell gene activation system identifies IGFBP2 as a mediator in the production of haematopoietic progenitors in vitro.
Por:
Petazzi, P, Ventura, T, Luongo, FP, McClafferty, H, May, A, Taylor, HA, Shipston, MJ, Romanò, N, Forrester, LM, Menendez, P and Fidanza, A
Publicada:
23 dic 2024
Ahead of Print:
23 dic 2024
Resumen:
A major challenge in the stem cell biology field is the ability to produce fully functional cells from induced pluripotent stem cells (iPSCs) that are a valuable resource for cell therapy, drug screening, and disease modelling. Here, we developed a novel inducible CRISPR-mediated activation strategy (iCRISPRa) to drive the expression of multiple endogenous transcription factors (TFs) important for in vitro cell fate and differentiation of iPSCs to haematopoietic progenitor cells. This work has identified a key role for IGFBP2 in developing haematopoietic progenitors. We first identified nine candidate TFs that we predicted to be involved in blood cell emergence during development, then generated tagged gRNAs directed to the transcriptional start site of these TFs that could also be detected during single-cell RNA sequencing (scRNAseq). iCRISPRa activation of these endogenous TFs resulted in a significant expansion of arterial-fated endothelial cells expressing high levels of IGFBP2, and our analysis indicated that IGFBP2 is involved in the remodelling of metabolic activity during in vitro endothelial to haematopoietic transition. As well as providing fundamental new insights into the mechanisms of haematopoietic differentiation, the broader applicability of iCRISPRa provides a valuable tool for studying dynamic processes in development and for recapitulating abnormal phenotypes characterised by ectopic activation of specific endogenous gene expression in a wide range of systems.
Filiaciones:
Petazzi, P:
Josep Carreras Leukemia Res Inst, Barcelona, Spain
Ventura, T:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
Luongo, FP:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
McClafferty, H:
Univ Edinburgh, Edinburgh Med Sch, Ctr Discovery Brain Sci, Biomed Sci, Edinburgh, Midlothian, Scotland
May, A:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
Taylor, HA:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
Shipston, MJ:
Univ Edinburgh, Edinburgh Med Sch, Ctr Discovery Brain Sci, Biomed Sci, Edinburgh, Midlothian, Scotland
Zhejiang Univ, Zhejiang Univ Univ Edinburgh Joint Inst, Sch Med, Haining, Peoples R China
Romanò, N:
Univ Edinburgh, Edinburgh Med Sch, Ctr Discovery Brain Sci, Biomed Sci, Edinburgh, Midlothian, Scotland
Zhejiang Univ, Zhejiang Univ Univ Edinburgh Joint Inst, Sch Med, Haining, Peoples R China
Forrester, LM:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
:
Josep Carreras Leukemia Res Inst, Barcelona, Spain
CIBER ONC, ISCIII, Barcelona, Spain
ICREA, Barcelona, Spain
Univ Barcelona, Sch Med, Dept Biomedicine, Barcelona, Spain
Inst Recerca St Joan de Deu PCCB SJD, Pediat Canc Ctr Barcelona, Barcelona, Spain
Fidanza, A:
Univ Edinburgh, Inst Regenerat & Repair, Ctr Regenerat Med, Edinburgh, Midlothian, Scotland
Univ Edinburgh, Edinburgh Med Sch, Biomed Sci, Edinburgh, Midlothian, Scotland
Green Submitted, gold
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