CRISPR-Cas9 interrogation of a putative fetal globin repressor in human erythroid cells

• DOI: 10.1371/journal.pone.0208237
• Publication Date: 2019-01-15T18:39:42Z
• Authors: Jennifer E. Chung; Wendy Magis; Jonathan Vu; Seok-Jin Heo; Kirmo Wartiovaara; Mark C. Walters; Ryo Kurita; Yukio Nakamura; Dario Boffelli; David I. K. Martin; Jacob E. Corn; Mark A. DeWitt
• Keywords: 0301 basic medicine; Genotype; Science; DISEASE; Cell Line; 03 medical and health sciences; Erythroid Cells; CRISPR-Associated Protein 9; Humans; gamma-Globins; Gene Silencing; HEMOGLOBIN; Fetal Hemoglobin; Sequence Deletion; Gene Editing; 0303 health sciences; Q; R; Hematopoietic Stem Cells; 16. Peace & justice; GENE; Up-Regulation; 3. Good health; LIFE; Repressor Proteins; Biomedicine; Phenotype; Medicine; BRII recipient: DeWitt; DNA, Intergenic; CRISPR-Cas Systems; Research Article

Sickle Cell Disease and ß-thalassemia, which are caused by defective or deficient adult ß-globin (HBB) respectively, the most common serious genetic blood diseases in world. Persistent expression of fetal ß-like globin, also known as 𝛾-globin, can ameliorate both disorders serving place a part hemoglobin tetramer (HbF). Here we use CRISPR-Cas9 gene editing to explore potential 𝛾-globin silencer region upstream δ-globin identified comparison naturally-occurring deletion mutations associated with up-regulated 𝛾-globin. We find that 1.7 kb consensus element select 350 bp sub-regions from bulk populations cells increases levels HbF. Screening individual sgRNAs one sub-region revealed three single guides expression. Deletion HUDEP-2 clonal sublines, colonies derived CD34+ hematopoietic stem/progenitor (HSPCs), does not cause significant up-regulation These data suggest is an autonomous silencer, thus itself suitable therapeutic target for treatment ß-hemoglobinopathies.