A Genome-wide CRISPR Death Screen Identifies Genes Essential for Oxidative Phosphorylation.
Oxidative phosphorylation (OXPHOS) is the major pathway for ATP production in humans. Deficiencies in OXPHOS can arise from mutations in either mitochondrial or nuclear genomes and comprise the largest collection of inborn errors of metabolism. At present we lack a complete catalog of human genes and pathways essential for OXPHOS. Here we introduce a genome-wide CRISPR "death screen" that actively selects dying cells to reveal human genes required for OXPHOS, inspired by the classic observation that human cells deficient in OXPHOS survive in glucose but die in galactose. We report 191 high-confidence hits essential for OXPHOS, including 72 underlying known OXPHOS diseases. Our screen reveals a functional module consisting of NGRN, WBSCR16, RPUSD3, RPUSD4, TRUB2, and FASTKD2 that regulates the mitochondrial 16S rRNA and intra-mitochondrial translation. Our work yields a rich catalog of genes required for OXPHOS and, more generally, demonstrates the power of death screening for functional genomic analysis.
Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Jason D
JD
Arroyo
Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Alexis A
AA
Jourdain
Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Sarah E
SE
Calvo
Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
Carmine A
CA
Ballarano
Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
John G
JG
Doench
Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
David E
DE
Root
Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address: vamsi@hms.harvard.edu.
Vamsi K
VK
Mootha
D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D005690:Galactose / Q000494:pharmacology; D050259:Genes, Mitochondrial; D016678:Genome; D005947:Glucose / Q000494:pharmacology; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D020014:K562 Cells; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D010085:Oxidative Phosphorylation; D010641:Phenotype; D014176:Protein Biosynthesis / Q000187:drug effects; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D015203:Reproducibility of Results
D012336:RNA, Ribosomal, 16S; D005947:Glucose; D005690:Galactose
16S RNA; CRISPR; Mitochondria; OXPHOS; death screen; mitochondrial ribosome; neugrin; pseudouridine; pseudouridylation; synthetic lethality
Cell. 2008 Jul 11;134(1):112-23
18614015
Genome Biol. 2014 Apr 10;15(4):R64
24721214
Nucleic Acids Res. 2016 Jan 4;44(D1):D1251-7
26450961
J Biol Chem. 1979 Apr 25;254(8):2669-76
429309
Genome Biol. 2014;15(12):554
25476604
Biochem Biophys Res Commun. 2000 Dec 20;279(2):526-33
11118320
Cell. 2012 Jun 8;149(6):1393-406
22658674
BMC Bioinformatics. 2009 Feb 03;10:48
19192299
J Exp Med. 1995 Nov 1;182(5):1545-56
7595224
J Mol Biol. 1997 Feb 21;266(2):246-68
9047361
Nature. 2007 Nov 29;450(7170):736-40
18046414
J Med Genet. 2012 Jan;49(1):10-5
22114105
Proc Natl Acad Sci U S A. 2008 Dec 23;105(51):20380-5
19091943
Semin Fetal Neonatal Med. 2011 Aug;16(4):197-204
21680271
JAMA. 2014 Jul 2;312(1):68-77
25058219
J Clin Invest. 2001 Oct;108(8):1167-74
11602624
Sci Am. 1997 Aug;277(2):40-7
9245840
Cell Rep. 2015 Feb 12;:null
25683715
Nat Rev Neurol. 2013 Aug;9(8):474-81
23817350
Cell. 2012 Mar 16;148(6):1145-59
22424226
Biochem Med Metab Biol. 1992 Oct;48(2):122-6
1329873
Nat Biotechnol. 2016 Feb;34(2):184-91
26780180
Nat Biotechnol. 2010 Mar;28(3):249-55
20160716
Am J Hum Genet. 2015 Nov 5;97(5):761-8
26522469
Nat Genet. 2002 Aug;31(4):400-4
12134146
Methods Cell Biol. 2007;80:571-88
17445714
Science. 1989 Oct 27;246(4929):500-3
2814477
RNA. 2015 Nov;21(11):1873-84
26370583
Elife. 2014 May 13;3:e02242
24843020
Am J Hum Genet. 2008 Sep;83(3):415-23
18771761
Mitochondrion. 2011 May;11(3):467-75
21292037
Cell. 2015 Dec 3;163(6):1515-26
26627737
Cell Rep. 2015 Feb 24;10(7):1110-21
25704814
Nature. 2012 Nov 15;491(7424):374-83
23151580