RED BLOOD CELLS

Domain-focused CRISPR screenidentifies HRI as a fetal hemoglobinregulator in human erythroid cellsJeremy D. Grevet1,2*, Xianjiang Lan1*, Nicole Hamagami1, Christopher R. Edwards1,Laavanya Sankaranarayanan1, Xinjun Ji2, Saurabh K. Bhardwaj1, Carolyne J. Face1,David F. Posocco1, Osheiza Abdulmalik1, Cheryl A. Keller3, Belinda Giardine3,Simone Sidoli4, Ben A. Garcia4, Stella T. Chou1, Stephen A. Liebhaber2,Ross C. Hardison3, Junwei Shi5†, Gerd A. Blobel1,2†

Increasing fetal hemoglobin (HbF) levels in adult red blood cells provides clinical benefitto patients with sickle cell disease and some forms of b-thalassemia. To identify potentiallydruggable HbF regulators in adult human erythroid cells, we employed a protein kinasedomain–focused CRISPR-Cas9–based genetic screen with a newly optimized single-guideRNA scaffold. The screen uncovered the heme-regulated inhibitor HRI (also known asEIF2AK1), an erythroid-specific kinase that controls protein translation, as an HbFrepressor. HRI depletion markedly increased HbF production in a specific manner andreduced sickling in cultured erythroid cells. Diminished expression of the HbF repressorBCL11A accounted in large part for the effects of HRI depletion. Taken together, theseresults suggest HRI as a potential therapeutic target for hemoglobinopathies.

The human b-globin gene cluster comprisesan embryonic [e-globin (HBE)], two fetal[g-globin (HBG1 andHBG2)], and two adult-type [d-globin and b-globin (HBDandHBB)]genes (1). Diseases affecting the adult-type

b-globin genes, such as sickle cell disease (SCD)and some types of b-thalassemia, manifest them-selves after birth following the transition fromfetal to adult hemoglobin production. The clin-ical course of these diseases is ameliorated whenfetal hemoglobin (HbF) levels are elevated be-cause of genetic causes or medical intervention(2). Genome-wide association studies have iden-tified three major loci that are linked to nat-ural variation in HbF levels, including that ofthe transcription factor BCL11A (1). Patients withheterozygous loss of BCL11A exhibit elevatedHbF levels throughout adult life (3), and BCL11Adepletion in cultured human erythroid precur-sors raises HbF production (4, 5). Loss of BCL11Ain engineered mice that express human sicklehemoglobin (HbS) stimulates fetal globin expres-sion and ameliorates the disease phenotype (6).Strategies to achieve long-term elevation of

HbF levels in patients include gene therapy toforce the expression of g-globin, as well as geneediting to impede BCL11A expression (7, 8). Cur-

rent clinical risks, costs, and availability limitsuch therapies to a fraction of the patient pop-ulation. The only FDA-approved drug to raiseHbF is hydroxyurea, which benefits many pa-tients but is limited in its efficacy (9). Hence,pharmacologic approaches to increase HbF levelsare needed.The transcription factors BCL11A and LRF

(ZBTB7A) and their coregulators mediate mostof g-globin transcriptional silencing in adult ery-throid cells (10). However, transcription factorsare inherently challenging to inhibit with smallmolecules. We carried out a CRISPR-Cas9 screento target protein kinases because, in principle,protein kinases are inherently controllable bysmall molecules (11). It has recently been shownthat targeting single-guide RNAs (sgRNAs) tofunctional protein domains can substantiallyimprove genetic screening efficiency, as both in-frame and frameshift mutations contribute togenerating hypomorphic alleles (12). We designeda library of sgRNAs targeting 482 kinase domains(6 sgRNAs per kinase domain and 50 nontar-geting sgRNAs as negative controls), which cov-ers almost all annotated kinases in the humangenome (11).Before performing the screen, we optimized

the Streptococcus pyogenes Cas9 sgRNA scaffoldfor on-target activity (see supplementary text fordetails). sgRNA2.1 emerged as the most efficientsgRNA in mutagenizing target loci in multiplecell lines, achieving up to 98% genome-editingefficiency as early as 4 days after introductioninto cells (figs. S1 and S2). We cloned the kinasedomain sgRNA library into the sgRNA2.1 scaf-fold and introduced it into HUDEP2 cells stablyexpressing Cas9 (Fig. 1A). HUDEP2 is an im-mortalized human cell line that can be inducedto undergo erythroid differentiation and pro-

duce high levels of adult-type and low levels offetal-type hemoglobins (13). We used anti-HbFfluorescence-activated cell sorting (FACS) toisolate the top 10% and bottom 10% of HbF-expressing cells (Fig. 1, A and B) and deep-sequenced sgRNAs as described previously(14). Nontargeting control sgRNAs were similarlydistributed across low- and high-HbF-expressionpopulations, indicating that the screen did notbias the enrichment in either direction (Fig. 1C).Heme-regulated inhibitor HRI (also known

as EIF2AK1) was the only kinase for which alltargeting sgRNAs were overrepresented in thehigh-HbF population (Fig. 1C). HRI is one offour kinases known to phosphorylate the proteintranslation initiation factor eIF2a (fig. S3) (15).Phosphorylation of eIF2a generally impedesmRNA translation but enables the translationof transcripts that include one or more upstreamopen reading frames in their 5′ untranslatedregions (15). HRI is enriched in erythroid cells,where it can be inhibited by its natural ligandheme (fig. S4, A and B). It is thought that thisregulation balances heme levels with produc-tion of globin chains, as excess of either of thesecan be detrimental (15). The other eIF2a kinasesare expressed at significantly lower levels inerythroid cells (fig. S4C). Though small moleculeshave been developed against other members ofthe eIF2a kinase family, effective HRI-specificinhibitors are lacking.We validated HRI as an HbF regulator by ex-

pressing all six “hit” sgRNAs (fig. S5) individual-ly in HUDEP2-Cas9 cells, along with an sgRNAagainst BCL11A as a benchmark and a nontar-geting sgRNA as a negative control. All six HRI-targeting sgRNAs increased the fraction of HbF+

cells (Fig. 2, A and B). We studied cells ex-pressing HRI sgRNAs 3 and 5 by Western blot-ting, which revealed that the sgRNAs achievedstrong depletion of HRI, resulting in decreasedeIF2a phosphorylation, and significantly increasedg-globin protein levels (Fig. 2C). There were nochanges in GATA1 protein levels, suggesting thatHRI loss did not impair cell maturation (Fig. 2C).eIF2a phosphorylation is thought to broadly

control protein translation, suggesting that HRIinhibition may lead to widespread changes inprotein levels and secondary changes in geneexpression. We measured protein abundances inundifferentiated and differentiated HRI-depletedHUDEP2 pools for sgRNAs 3 and 5 by replicatewhole-cell mass spectrometry studies (fig. S6).Notably, protein amounts were changed rela-tively little in HRI-depleted differentiated HUDEP2cells (Fig. 2D and fig. S7), and g-globin was oneof the most induced proteins (Fig. 2D, fig. S7A,and table S1). We did observe a modest risein global protein levels, perhaps because of anonspecific augmentation in protein translationresulting from lower eIF2a phosphorylation(fig. S7, A and B). Whole-cell mass spectrometrydetected the ~2000 and ~2500 most abundantproteins in differentiated and undifferentiatedsamples, respectively. Thus, it is possible thatsignificant changes were missed among low-abundance proteins. Nevertheless, overall the

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Grevet et al., Science 361, 285–290 (2018) 20 July 2018 1 of 6

1Division of Hematology, The Children’s Hospital ofPhiladelphia, Philadelphia, PA 19104, USA. 2Department ofGenetics, Perelman School of Medicine, University ofPennsylvania, Philadelphia, PA 19104, USA. 3Department ofBiochemistry and Molecular Biology, Pennsylvania StateUniversity, University Park, PA 16802, USA. 4Department ofBiochemistry and Biophysics, Perelman School of Medicine,University of Pennsylvania, Philadelphia, PA 19104, USA.5Department of Cancer Biology, Perelman School of Medicine,University of Pennsylvania, Philadelphia, PA 19104, USA.*These authors contributed equally to this work.†Corresponding author. Email: jushi@upenn.edu (J.S.); blobel@email.chop.edu (G.A.B.)

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effects of HRI depletion were unexpectedlyspecific for g-globin.In parallel, we carried out replicate RNA

sequencing (RNA-seq) experiments (fig. S8A)in differentiated HRI-depleted HUDEP2 poolsfor sgRNAs 3 and 5 and found that global tran-script distributions were relatively unperturbed(Fig. 2E). g-Globin mRNA stood out as the mostsignificantly induced in differentiated cells(Fig. 2E, fig. S8B, and table S1). Reverse tran-scription quantitative polymerase chain reaction(RT-qPCR) experiments confirmed the effects ong-globin mRNA and primary transcripts, suggest-ing that HRI depletion increases g-globin tran-scription (fig. S9, A and B). We did not observeany noticeable changes in a-globin, ALAS2,BAND3, and GATA1 mRNAs (fig. S9C), suggest-ing that erythroid differentiation is essentiallynormal in HRI-deficient cells. These resultswere further validated in two independent clonalHRI-depleted cell lines generated from the HRIsgRNA 5 pool (fig. S10).

HRI’s role as an HbF repressor was furthertested in primary erythroblasts derived from athree-phase human CD34+ culture system (seematerials and methods). We depleted HRI withtwo independent short hairpin RNAs (shRNAs).For comparison, we used the stronger shRNAagainst LRF among two previously described(10). We verified transduction levels by greenfluorescent protein (GFP) flow cytometry (fig.S11A). Both HRI shRNAs significantly increasedthe proportion of HbF+ cells (Fig. 3, A and B).The proportion of g-globin mRNA as a percentageof total globin transcripts was also significantlyincreased upon HRI knockdown (Fig. 3C).RNA-seq in cells with either shRNA demon-

strated that g-globin mRNAwas among the moststrongly increased (Fig. 3D), similar to the re-sults in HUDEP2 cells, with global transcriptdistributions for HRI knockdown samples beingstrongly correlatedwith those from the scrambledshRNA (Fig. 3D). Similar elevation of HbF wasdetected at the protein level, as measured by

high-performance liquid chromatography (HPLC)(Fig. 3E; note donor-to-donor variation in thehuman culture system). HRI depletion did notappear to delay the maturation of these cells, asevidenced by similar transcript levels for matu-ration markers (Fig. 3D) and cell surface mark-ers CD71 and CD235a (fig. S11B) and similar cellmorphologies (fig. S11C).Elevated HbF levels counteract sickling of

cells expressing HbS (1). To assess whether HRIdepletion elicits antisickling effects, we knockeddown HRI in CD34+ cells obtained from pa-tients with SCD (fig. S12A). This led to increasesin g-globin levels comparable to those in theCD34+ cultures from healthy donors mentionedabove (fig. S12, B and C) and, as before, did notseem to affect erythroid maturation (fig. S12D).When grown under low oxygen tension, cells ex-pressing HRI shRNAs were less prone to sicklingthan controls (Fig. 3F), suggesting that HRI de-pletion may achieve therapeutically relevant lev-els of HbF.

Grevet et al., Science 361, 285–290 (2018) 20 July 2018 2 of 6

Fig. 1. A kinase domain–focused CRISPR-Cas9 screen identifiesHRI as a novel fetal globin repressor. (A) Experimental outline. Seematerials and methods for a full description. Oligo, oligonucleotide; MOI,multiplicity of infection; gDNA, genomic DNA. (B) Representative HbF

FACS gating strategy for sorting high-HbF and low-HbF populations forscreens. FSC, forward scatter; K, thousand. (C) sgRNA representation inlow-HbF versus high-HbF populations as log2-transformed normalizedread counts.

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BCL11A and LRF are the major direct re-pressors of g-globin transcription (10), suggest-ing that the effects of HRI may converge onone or both of these factors, as they are expressedduring overlapping time intervals of erythroidmaturation (fig. S13). We examined BCL11A andLRF protein levels by Western blotting with ex-tracts from CD34+ cell-derived erythroblasts ex-pressing HRI shRNAs. Notably, BCL11A but notLRF was strongly depleted in both HRI knock-down samples (Fig. 4A). Western blots confirmedthe loss of HRI, the resulting reduction in phospho-eIF2a, and the increase in g-globin (Fig. 4A). GATA1was unaffected, in agreement with ostensiblynormal cell maturation under conditions of HRIdepletion.We tested whether HRI controls BCL11A

levels directly by governing its translation orindirectly—for example, by controlling the trans-lation of a nuclear factor that promotes BCL11Atranscription. Both BCL11A mRNA and primary

transcript levels were markedly reduced in HRI-depleted cells, indicating that most of the BCL11Aloss is a result of transcription inhibition in HRI-depleted primary erythroid cells, HUDEP2 pools,and clonal lines (Fig. 4A and fig. S14). To mea-sure a possible contribution of direct translationalcontrol of BCL11A mRNA by HRI, we performedpolysome profiling. HRI depletion modestly in-creased overall translation efficiency, as poly-somal fractions were slightly more abundantthan monosomes (fig. S15A). Moreover, in HRI-depleted cells, BCL11A mRNA levels were signif-icantly reduced across all polysomal fractions,consistent with an overall loss in mRNA levels(fig. S15B). To better visualize distribution acrossfractions, results were plotted as a percentageof total BCL11A mRNA, which revealed littleshift in the distribution pattern (fig. S15C), sug-gesting that BCL11A mRNA translation playsonly a minor if any role in response to HRI loss.a-Globin mRNAs tended to shift slightly to the

larger polysome fractions (to the right), con-sistent with a subtle increase in their translation(fig. S15C).To assess to what extent BCL11A loss accounts

for the effects of HRI depletion, BCL11A expres-sion was restored in HRI-depleted HUDEP2clonal cell lines. This reinstated g-globin repres-sion by 65 to 80% (Fig. 4B and fig. S16), suggest-ing that BCL11A is amajor HRI effector. Of note,depleting HRI in primary definitive (adult-type)erythroid cells from murine fetal livers did notreduce BCL11A levels (fig. S17), suggesting thatcertain pathways controlled by HRI may be spe-cies specific. In sum, BCL11A emerges as an HRItarget in the control of HbF production in humanadult erythroid cells.As a proof of principle, we assessed whether

HRI’s effects on HbF can be amplified with phar-macologic HbF inducers. Pomalidomide has beenrecently shown to induce HbF in part by reducingBCL11A transcription (16). We tested the effects

Grevet et al., Science 361, 285–290 (2018) 20 July 2018 3 of 6

Fig. 2. HRI depletion elevates g-globin in HUDEP2 cells. (A) Rep-resentative HbF FACS for HUDEP2-Cas9 sgRNA pools. (B) Summary of HbFflow cytometry of cells expressing indicated sgRNAs. Data are means ± SDfrom biological replicates (n = 2). (C) Western blots with antibodies againstthe indicated proteins in cell pools expressing sgRNAs as indicated. Diff,differentiation. (D) Mass spectrometry for the HRI sgRNA 5 pool from

biological replicates (n = 3). Data are log2 mean-normalized proteinabundances in cells with HRI sgRNA 5 and nontargeting sgRNA. r is thePearson correlation coefficient. HBZ, z-globin; HBA1/2, a-globins 1 and 2.(E) RNA-seq of cell pools with HRI sgRNA 5. Data are log2 numbers offragments per kilobase of transcript per million mapped reads (FPKM)averaged from biological replicates (n = 2).

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Fig. 3. HRI depletion elevates g-globin in primary erythroid CD34+

cells. (A) Representative HbF flow cytometry on day 15 of CD34+

erythroid differentiation. SC, sickle cell. (B) Summary of HbF flowcytometry experiments. Error bars represent SEM from threeindependent donors (*P < 0.05, **P < 0.01 from unpaired Studentt tests). (C) g-Globin mRNA as a fraction of g-globin plus b-globin byRT-qPCR on day 13. Error bars represent SEM from three independentdonors (*P < 0.05, **P < 0.01 from unpaired Student t tests). (D) RNA-seq

for CD34+ cells on day 13. r is the Pearson correlation coefficient.Data were obtained from one patient donor. (E) HPLC analysisof samples from cells expressing annotated shRNAs at day 15 ofdifferentiation. HbA, hemoglobin A (adult form). Error bars representSD from two independent donors. (F) Images of sickle cell patient–derived erythroid cultures. Arrowheads mark cells with sickle-likemorphology. The bar graph summarizes blind counts of sickle-shapedcells in three fields from one patient with SCD.

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of pomalidomide treatment on two HRI-depletedHUDEP2 populations and control HUDEP2cells. For comparison, we also included HUDEP2cells in which BCL11A transcription was reducedby an sgRNA targeting the erythroid BCL11Aenhancer. These combinations did not impaircell viability or maturation (fig. S18, A to D). Allconditions alone elevated the fraction of HbF+

cells and increased g-globin transcription as ex-pected (Fig. 4C and fig. S18, B, C, and E). Thestrongest cooperativity was observed when HRIdepletion was combined with pomalidomide.Using an improved CRISPR-Cas9 domain–

focused screening approach, we identified theerythroid-specific kinase HRI as a potentiallydruggable target that is involved in HbF silenc-ing. Our findings contrast with previous reportsshowing that stabilization of eIF2a phosphoryl-ation with the drug salubrinal can stimulateHbF synthesis in cultured human erythroid

cells (17, 18). Salubrinal acts downstream ofHRI, and the mechanism underlying the induc-tion of HbF remains to be fully elucidated.Hematopoietic stress can be associated withincreased HbF levels. However, we believe thatthe effects of HRI depletion are not due to stressbecause HRI depletion seemingly has little ad-verse effect on cell viability and cell maturation,as judged by cell morphological, surface pheno-typing, transcriptome, and proteome analyses.Moreover, silencing of HbF in HRI-depleted cellsis largely reversed upon BCL11A reexpression,consistent with a specific function of HRI.Mice in which the HRI gene has been knocked

out appear for the most part normal and havenormal hematological indices (19). This findingis in agreement with our results in human ery-throid cell cultures and suggests that HRI lossis generally well tolerated. However, HRI micedisplayed impaired adaptation to erythropoietic

stress induced by iron deficiency and had ex-acerbated protoporphyria and thalassemia phe-notypes (20, 21). It remains to be seen whetherHRI inhibition in SCD patients would elevateHbF levels sufficiently to improve outcomes. HRIinhibition elevatedHbF levels to a point at whichit reduced cell sickling in culture, suggesting thatpharmacologic HRI inhibitors may provide clin-ical benefit in SCD patients. Moreover, in light ofour results, combining HRI inhibition with anadditional pharmacologic HbF inducer may im-prove the therapeutic index.

REFERENCES AND NOTES

1. V. G. Sankaran, S. H. Orkin, Cold Spring Harb. Perspect. Med. 3,a011643 (2013).

2. O. S. Platt et al., N. Engl. J. Med. 330, 1639–1644 (1994).3. A. Basak et al., J. Clin. Invest. 125, 2363–2368 (2015).4. V. G. Sankaran et al., Science 322, 1839–1842 (2008).5. V. G. Sankaran et al., Nature 460, 1093–1097 (2009).6. J. Xu et al., Science 334, 993–996 (2011).

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Fig. 4. HRI regulates BCL11A levels. (A) (Left) Western blotwith the indicated antibodies in uninfected cells or cellsinfected with virus expressing scrambled or HRI shRNAson day 15 of differentiation. (Right) BCL11A mRNA (n = 4independent donors) and primary (1°) transcript (n = 3independent donors) levels in HRI knockdown cells as afraction of those in SC shRNA–expressing cells. Error barsrepresent SD from biological replicates. (B) BCL11Acomplementary DNA (cDNA) rescue in HRI-depleted HUDEP2clonal cell lines. Data show RT-qPCR for g-globin as afraction of g-globin plus b-globin. Error bars indicate SD fromtwo biological replicates (C) HbF flow cytometry of indicatedHUDEP2 sgRNA pools grown in the presence or absenceof pomalidomide. Error bars indicate SEM of threebiological replicates.

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7. A. Perumbeti et al., Blood 114, 1174–1185 (2009).8. D. E. Bauer et al., Science 342, 253–257 (2013).9. S. M. Bakanay et al., Blood 105, 545–547 (2005).10. T. Masuda et al., Science 351, 285–289 (2016).11. G. Manning, D. B. Whyte, R. Martinez, T. Hunter,

S. Sudarsanam, Science 298, 1912–1934 (2002).12. J. Shi et al., Nat. Biotechnol. 33, 661–667 (2015).13. R. Kurita et al., PLOS ONE 8, e59890–e15 (2013).14. M. C. Canver et al., Nature 527, 192–197 (2015).15. J.-J. Chen, Curr. Opin. Hematol. 21, 172–178 (2014).16. B. M. Dulmovits et al., Blood 127, 1481–1492 (2016).17. C. K. Hahn, C. H. Lowrey, Blood 122, 477–485 (2013).18. C. K. Hahn, C. H. Lowrey, Blood 124, 2730–2734 (2014).19. A. P. Han et al., EMBO J. 20, 6909–6918 (2001).20. S. Liu et al., Br. J. Haematol. 143, 129–137 (2008).21. S. Liu et al., Haematologica 93, 753–756 (2008).

ACKNOWLEDGMENTS

This paper is dedicated to the memory of Günter Blobel and hisboundless generosity and ability to inspire. We thank C. Vakoc forproviding the kinase-focused sgRNA library and members of the

lab for helpful discussions. We thank the Children’s Hospital ofPhiladelphia flow cytometry core for experimental assistance andR. Kurita and Y. Nakamura for contributing the HUDEP2 cells. Funding:J.D.G. was supported by T32GM008216 and 1F30DK107055-01;B.A.G. by W81XWH-113-1-0426, R01-GM110174, and R01-AI118891;R.C.H. by R56DK065806 and U54HG006998; S.A.L. byR01-HL065449; J.S. by Cold Spring Harbor Laboratory sponsoredresearch; and G.A.B. by 5R01HL119479 and 5R37DK058044.Author contributions: J.D.G., J.S., and G.A.B. conceived of the studyand designed the experiments. J.S. designed and executed thesgRNA and Cas9 plasmid cloning, sgRNA scaffold optimization,sgRNA library construction, and viral packaging. J.D.G. executed theHUDEP2-Cas9 editing optimization and CRISPR screen. J.D.G., J.S.,X.L., N.H., C.R.E., L.S., S.K.B., C.J.F., D.F.P., O.A., and S.T.C. carried outexperiments. S.S. and B.A.G. performed mass spectrometry anddownstream analysis. C.A.K., B.G., and R.C.H. executed RNA-seq andread alignment. J.D.G., X.J., and S.A.L. designed and performed thepolysome profiling experiments. J.D.G., J.S., and G.A.B. wrote themanuscript. Competing interests: J.D.G., J.S., and G.A.B. areinventors on a patent (Patent Cooperation Treaty patent applicationPCT/US18/15918) submitted by The Children’s Hospital of

Philadelphia that covers the therapeutic targeting of HRI forhemoglobinopathies. Data and materials availability: Allplasmids and reagents described in this study are available tothe scientific community upon request to J.S. and G.A.B. Massspectrometry data are deposited at https://chorusproject.org(project 1488), and RNA-seq data are available at the NCBI GeneExpression Omnibus (GEO accession number GSE115687).

SUPPLEMENTARY MATERIALS

www.sciencemag.org/content/361/6399/285/suppl/DC1Materials and MethodsSupplementary TextFigs. S1 to S18Tables S1 to S7References (22–30)Data S1 to S5

22 July 2017; resubmitted 15 April 2018Accepted 13 June 201810.1126/science.aao0932

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erythroid cellsDomain-focused CRISPR screen identifies HRI as a fetal hemoglobin regulator in human

Sidoli, Ben A. Garcia, Stella T. Chou, Stephen A. Liebhaber, Ross C. Hardison, Junwei Shi and Gerd A. BlobelSaurabh K. Bhardwaj, Carolyne J. Face, David F. Posocco, Osheiza Abdulmalik, Cheryl A. Keller, Belinda Giardine, Simone Jeremy D. Grevet, Xianjiang Lan, Nicole Hamagami, Christopher R. Edwards, Laavanya Sankaranarayanan, Xinjun Ji,

DOI: 10.1126/science.aao0932 (6399), 285-290.361Science 

, this issue p. 285Sciencehuman RBCs. Thus, HRI may be a therapeutic target for sickle cell disease and other hemoglobin disorders.kinase (HRI). Depleting the kinase in RBCs led to an increase in fetal hemoglobin levels and reduced sickling of cultured

α performed a CRISPR-Cas9 screen for regulators of fetal hemoglobin in RBCs and identified heme-regulated eIF2et al.Grevetthat involves abnormal hemoglobin. Current treatments entail modulating the level of fetal hemoglobin expression.

Hemoglobin in red blood cells (RBCs) carries oxygen to the tissues. Sickle cell disease is an inherited conditionA CRISPR screen for RBC regulators

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