CRISPR-Cas9 interrogation of a putative fetal globin repressor in human erythroid cells.
CRISPR-Associated Protein 9
/ metabolism
CRISPR-Cas Systems
/ genetics
Cell Line
DNA, Intergenic
/ genetics
Erythroid Cells
/ metabolism
Fetal Hemoglobin
/ metabolism
Gene Editing
Gene Silencing
Genotype
Hematopoietic Stem Cells
/ metabolism
Humans
Phenotype
Repressor Proteins
/ metabolism
Sequence Deletion
/ genetics
Up-Regulation
/ genetics
gamma-Globins
/ genetics
Journal
PloS one
ISSN: 1932-6203
Titre abrégé: PLoS One
Pays: United States
ID NLM: 101285081
Informations de publication
Date de publication:
2019
2019
Historique:
received:
19
07
2018
accepted:
14
11
2018
entrez:
16
1
2019
pubmed:
16
1
2019
medline:
26
9
2019
Statut:
epublish
Résumé
Sickle Cell Disease and ß-thalassemia, which are caused by defective or deficient adult ß-globin (HBB) respectively, are the most common serious genetic blood diseases in the world. Persistent expression of the fetal ß-like globin, also known as 𝛾-globin, can ameliorate both disorders by serving in place of the adult ß-globin as a part of the fetal hemoglobin tetramer (HbF). Here we use CRISPR-Cas9 gene editing to explore a potential 𝛾-globin silencer region upstream of the δ-globin gene identified by comparison of naturally-occurring deletion mutations associated with up-regulated 𝛾-globin. We find that deletion of a 1.7 kb consensus element or select 350 bp sub-regions from bulk populations of cells increases levels of HbF. Screening of individual sgRNAs in one sub-region revealed three single guides that caused increases in 𝛾-globin expression. Deletion of the 1.7 kb region in HUDEP-2 clonal sublines, and in colonies derived from CD34+ hematopoietic stem/progenitor cells (HSPCs), does not cause significant up-regulation of 𝛾-globin. These data suggest that the 1.7 kb region is not an autonomous 𝛾-globin silencer, and thus by itself is not a suitable therapeutic target for gene editing treatment of ß-hemoglobinopathies.
Identifiants
pubmed: 30645582
doi: 10.1371/journal.pone.0208237
pii: PONE-D-18-21372
pmc: PMC6333401
doi:
Substances chimiques
DNA, Intergenic
0
Repressor Proteins
0
gamma-Globins
0
Fetal Hemoglobin
9034-63-3
CRISPR-Associated Protein 9
EC 3.1.-
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e0208237Subventions
Organisme : NIH HHS
ID : S10 OD018174
Pays : United States
Déclaration de conflit d'intérêts
The authors have declared that no competing interests exist.
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