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Nature. 2017 Apr 12;544(7649):235-239. doi: 10.1038/nature22034.

Human knockouts and phenotypic analysis in a cohort with a high rate of consanguinity.

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Department of Biostatistics and Epidemiology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Center for Non-Communicable Diseases, Karachi, Pakistan.
Center for Genomic Medicine, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, Massachusetts, USA.
Broad Institute of Harvard and MIT, Cambridge, Massachusetts, USA.
Analytic and Translational Genetics Unit, Department of Medicine, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts, USA.
Institute for Translational Medicine and Therapeutics, Department of Genetics, and Department of Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Samsung Advanced Institute for Health Sciences and Technology (SAIHST), Sungkyunkwan University, Samsung Medical Center, Seoul, Korea.
Division of Genetics and Genomics, Boston Children's Hospital, Boston, Massachusetts, USA.
Faisalabad Institute of Cardiology, Faisalabad, Pakistan.
National Institute of Cardiovascular Disorders, Karachi, Pakistan.
Punjab Institute of Cardiology, Lahore, Pakistan.
Karachi Institute of Heart Diseases, Karachi, Pakistan.
Red Crescent Institute of Cardiology, Hyderabad, Pakistan.
The Civil Hospital, Karachi, Pakistan.
Liaquat National Hospital, Karachi, Pakistan.
Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA.
The Charles Bronfman Institute for Personalized Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, USA.
Children's Hospital Oakland Research Institute, Oakland, California, USA.
MRC/BHF Cardiovascular Epidemiology Unit, Department of Public Health and Primary Care, University of Cambridge, UK.
Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK.
Department of Human Genetics, University of Pennsylvania, USA.


A major goal of biomedicine is to understand the function of every gene in the human genome. Loss-of-function mutations can disrupt both copies of a given gene in humans and phenotypic analysis of such 'human knockouts' can provide insight into gene function. Consanguineous unions are more likely to result in offspring carrying homozygous loss-of-function mutations. In Pakistan, consanguinity rates are notably high. Here we sequence the protein-coding regions of 10,503 adult participants in the Pakistan Risk of Myocardial Infarction Study (PROMIS), designed to understand the determinants of cardiometabolic diseases in individuals from South Asia. We identified individuals carrying homozygous predicted loss-of-function (pLoF) mutations, and performed phenotypic analysis involving more than 200 biochemical and disease traits. We enumerated 49,138 rare (<1% minor allele frequency) pLoF mutations. These pLoF mutations are estimated to knock out 1,317 genes, each in at least one participant. Homozygosity for pLoF mutations at PLA2G7 was associated with absent enzymatic activity of soluble lipoprotein-associated phospholipase A2; at CYP2F1, with higher plasma interleukin-8 concentrations; at TREH, with lower concentrations of apoB-containing lipoprotein subfractions; at either A3GALT2 or NRG4, with markedly reduced plasma insulin C-peptide concentrations; and at SLC9A3R1, with mediators of calcium and phosphate signalling. Heterozygous deficiency of APOC3 has been shown to protect against coronary heart disease; we identified APOC3 homozygous pLoF carriers in our cohort. We recruited these human knockouts and challenged them with an oral fat load. Compared with family members lacking the mutation, individuals with APOC3 knocked out displayed marked blunting of the usual post-prandial rise in plasma triglycerides. Overall, these observations provide a roadmap for a 'human knockout project', a systematic effort to understand the phenotypic consequences of complete disruption of genes in humans.

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