Abstract
Developmental silencing of fetal globins serves as both a paradigm of spatiotemporal gene regulation and an opportunity for therapeutic intervention of β-hemoglobinopathy. The nucleosome remodeling and deacetylase (NuRD) chromatin complex participates in γ-globin repression. We used pooled CRISPR screening to disrupt NuRD protein coding sequences comprehensively in human adult erythroid precursors. Essential for fetal hemoglobin (HbF) control is a non-redundant subcomplex of NuRD protein family paralogs, whose composition we corroborated by affinity chromatography and proximity labeling mass spectrometry proteomics. Mapping top functional guide RNAs identified key protein interfaces where in-frame alleles resulted in loss-of-function due to destabilization or altered function of subunits. We ascertained mutations of CHD4 that dissociate its requirement for cell fitness from HbF repression in both primary human erythroid precursors and transgenic mice. Finally we demonstrated that sequestering CHD4 from NuRD phenocopied these mutations. These results indicate a generalizable approach to discover protein complex features amenable to rational biochemical targeting.
| Original language | English |
|---|---|
| Journal | Nature genetics |
| DOIs | |
| Publication status | Published - Jan 1 2019 |
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All Science Journal Classification (ASJC) codes
- Genetics
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Rational targeting of a NuRD subcomplex guided by comprehensive in situ mutagenesis. / Sher, Falak; Hossain, Mir; Seruggia, Davide; Schoonenberg, Vivien A.C.; Yao, Qiuming; Cifani, Paolo; Dassama, Laura M.K.; Cole, Mitchel A.; Ren, Chunyan; Vinjamur, Divya S.; Macias-Trevino, Claudio; Luk, Kevin; McGuckin, Connor; Schupp, Patrick G.; Canver, Matthew C.; Kurita, Ryo; Nakamura, Yukio; Fujiwara, Yuko; Wolfe, Scot A.; Pinello, Luca; Maeda, Takahiro; Kentsis, Alex; Orkin, Stuart H.; Bauer, Daniel E.
In: Nature genetics, 01.01.2019.Research output: Contribution to journal › Article
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TY - JOUR
T1 - Rational targeting of a NuRD subcomplex guided by comprehensive in situ mutagenesis
AU - Sher, Falak
AU - Hossain, Mir
AU - Seruggia, Davide
AU - Schoonenberg, Vivien A.C.
AU - Yao, Qiuming
AU - Cifani, Paolo
AU - Dassama, Laura M.K.
AU - Cole, Mitchel A.
AU - Ren, Chunyan
AU - Vinjamur, Divya S.
AU - Macias-Trevino, Claudio
AU - Luk, Kevin
AU - McGuckin, Connor
AU - Schupp, Patrick G.
AU - Canver, Matthew C.
AU - Kurita, Ryo
AU - Nakamura, Yukio
AU - Fujiwara, Yuko
AU - Wolfe, Scot A.
AU - Pinello, Luca
AU - Maeda, Takahiro
AU - Kentsis, Alex
AU - Orkin, Stuart H.
AU - Bauer, Daniel E.
PY - 2019/1/1
Y1 - 2019/1/1
N2 - Developmental silencing of fetal globins serves as both a paradigm of spatiotemporal gene regulation and an opportunity for therapeutic intervention of β-hemoglobinopathy. The nucleosome remodeling and deacetylase (NuRD) chromatin complex participates in γ-globin repression. We used pooled CRISPR screening to disrupt NuRD protein coding sequences comprehensively in human adult erythroid precursors. Essential for fetal hemoglobin (HbF) control is a non-redundant subcomplex of NuRD protein family paralogs, whose composition we corroborated by affinity chromatography and proximity labeling mass spectrometry proteomics. Mapping top functional guide RNAs identified key protein interfaces where in-frame alleles resulted in loss-of-function due to destabilization or altered function of subunits. We ascertained mutations of CHD4 that dissociate its requirement for cell fitness from HbF repression in both primary human erythroid precursors and transgenic mice. Finally we demonstrated that sequestering CHD4 from NuRD phenocopied these mutations. These results indicate a generalizable approach to discover protein complex features amenable to rational biochemical targeting.
AB - Developmental silencing of fetal globins serves as both a paradigm of spatiotemporal gene regulation and an opportunity for therapeutic intervention of β-hemoglobinopathy. The nucleosome remodeling and deacetylase (NuRD) chromatin complex participates in γ-globin repression. We used pooled CRISPR screening to disrupt NuRD protein coding sequences comprehensively in human adult erythroid precursors. Essential for fetal hemoglobin (HbF) control is a non-redundant subcomplex of NuRD protein family paralogs, whose composition we corroborated by affinity chromatography and proximity labeling mass spectrometry proteomics. Mapping top functional guide RNAs identified key protein interfaces where in-frame alleles resulted in loss-of-function due to destabilization or altered function of subunits. We ascertained mutations of CHD4 that dissociate its requirement for cell fitness from HbF repression in both primary human erythroid precursors and transgenic mice. Finally we demonstrated that sequestering CHD4 from NuRD phenocopied these mutations. These results indicate a generalizable approach to discover protein complex features amenable to rational biochemical targeting.
UR - http://www.scopus.com/inward/record.url?scp=85068316955&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85068316955&partnerID=8YFLogxK
U2 - 10.1038/s41588-019-0453-4
DO - 10.1038/s41588-019-0453-4
M3 - Article
C2 - 31253978
AN - SCOPUS:85068316955
JO - Nature Genetics
JF - Nature Genetics
SN - 1061-4036
ER -