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Refining the role of de novo protein-truncating variants in neurodevelopmental disorders by using population reference samples


Recent research has uncovered an important role for de novo variation in neurodevelopmental disorders. Using aggregated data from 9,246 families with autism spectrum disorder, intellectual disability, or developmental delay, we found that 1/3 of de novo variants are independently present as standing variation in the Exome Aggregation Consortium's cohort of 60,706 adults, and these de novo variants do not contribute to neurodevelopmental risk. We further used a loss-of-function (LoF)-intolerance metric, pLI, to identify a subset of LoF-intolerant genes containing the observed signal of associated de novo protein-truncating variants (PTVs) in neurodevelopmental disorders. LoF-intolerant genes also carry a modest excess of inherited PTVs, although the strongest de novo–affected genes contribute little to this excess, thus suggesting that the excess of inherited risk resides in lower-penetrant genes. These findings illustrate the importance of population-based reference cohorts for the interpretation of candidate pathogenic variants, even for analyses of complex diseases and de novo variation.

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Figure 1
Figure 2: Properties of class 2 de novo variants.
Figure 3: Partitioning the rate of de novo variants per exome on the basis of class 1, class 2, and pLI.
Figure 4: Phenotypic associations for ASD de novo PTVs.


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We thank all of the members of the ATGU and the laboratory of D.P.W. for assistance in this endeavor. We thank the families who took part in the Simons Simplex Collection study and the Simons Variation in Individuals Project, as well as the clinicians who collected data at each of the study sites. The authors would like to thank the Exome Aggregation Consortium and the groups that provided exome variant data for comparison. A full list of contributing groups can be found on the ExAC website (see URLs). We also greatly thank A. Byrnes, R. Fine, D. Fronk, A. Martin, C. Nichols, N. Radd, K. Satterstrom, and E. Wigdor for their insightful contributions. Finally, we acknowledge G.A. Barnard for inspiring us to write in a more conversational tone similar to that in his seminal 1947 paper (Biometrika 34, 123–138, 1947). This work was supported by NIH grants U01MH100233, U01MH100209, U01MH100229, and U01MH100239 to the Autism Sequencing Consortium (ASC), and R56 MH097849 and R01 MH097849 to the Population-based Autism Genetics and Environment Study (PAGES). M.J.D., J.A.K., and K.E.S. were supported by grants from the Simons Foundation Autism Research Initiative (SFARI 342292 and a subaward from the Simons Center for the Social Brain at MIT). M.L. and D.G.M.'s work on the ExAC project was funded by U54DK105566 and R01 GM104371 from the National Institutes of Health. K.S. was funded by T32 HG002295/HG/NHGRI. E.B.R. was funded by National Institutes of Mental Health Grant 1K01MH099286 and NARSAD Young Investigator grant 22379.

Author information




J.A.K. and E.B.R. performed the analyses. J.A.K., D.P.H., E.B.R., and M.J.D. designed the experiment. J.A.K. and K.S. wrote the code. D.P.W., E.B.R., and M.J.D. supervised the research. J.A.K. and M.J.D. wrote the paper. J.A.K., K.E.S., D.P.H., S.J.S., M.L., K.J.K., D.G.M., and J.D.B. generated data. J.A.K., K.E.S., D.P.H., D.J.C., B.D., K.R., B.M.N., D.G.M., D.P.W., E.B.R., and M.J.D. contributed to analysis concepts and methods. J.A.K. was responsible for the remainder. All authors revised and approved the final manuscript.

Corresponding author

Correspondence to Mark J Daly.

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Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Recurrence rate is a function of allele frequency and reference-population size.

Expected number of discovered class 2 de novo variants by size of the reference dataset, partitioned based on the number of copies of the variant currently present in ExAC. The number of de novo variants found in the standing population is a function of the sample size of the reference dataset and the current estimated allele count (AC).

Supplementary Figure 2 Recurrence rate for the nonpsychiatric version of ExAC.

The proportion of de novo variants across each cohort split between class 1 (left) and class 2 (right) when using the non-psychiatric version of ExAC (See Figure 2A for the results using the full version of ExAC). Removing the 15,330 exomes from the psychiatric cohorts did not change the enrichment of CpG variants among class 2nonpsych de novo variants (P < 10-20). Error bars represent 95% confidence intervals. ID/DD, intellectual disability / developmental delay; ASD, autism spectrum disorder.

Supplementary Figure 3 Partitioning the rate of de novo variants per exome on the basis of class 1, class 2, and pLI, by using the nonpsychiatric version of ExAC.

Within each grouping, the rate is shown for ID/DD (left), ASD (middle), and unaffected ASD siblings (right) with the number of individuals labeled in the legend. (a) Rate of de novo synonymous variants per exome partitioned into class 2nonpsych (middle) and class 1nonpsych (right). No significant difference was observed for any grouping of de novo synonymous variants. (b) Rate of de novo PTVs per exome partitioned into class 2nonpsych (middle) and class 1nonpsych (right). Only class 1nonpsych de novo PTVs in ID/DD and ASD show association when compared to unaffected ASD siblings. (c) Rate of de novo PTVs partitioned into class 1nonpsych de novo PTVs in pLI ≥0.9 genes (right), and the class 1nonpsych de novo PTVs in pLI <0.9 genes (middle). For all such analyses, the rate ratio and significance were calculated by comparing the rate for ID/DD and ASD to the rate in unaffected ASD siblings using a two-sided Poisson exact test for synonymous variants and one-sided for the remainder (Supplemental Note). Error bars represent 95% confidence intervals throughout (a) – (c). ID/DD, intellectual disability / developmental delay; ASD, autism spectrum disorder; PTV, protein truncating variant; pLI, probability of loss-of-function intolerance; NS, not significant.

Supplementary Figure 4 Recalibrating VQSLOD for transmission of singleton synonymous variants.

Singleton synonymous variants discovered in the joint called set was ordered by VQSLOD in descending order (i.e. higher confident variants first) and then binned into percentiles. The red line marks the GATK-defined VQSR cut off (-1.49) and the blue line marks where VQSR was moved (-1.724) to achieve 1:1 transmission rate of synonymous variants.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4, Supplementary Tables 3–5, 7–21, 23–25, 27–30 and 32–34, and Supplementary Note (PDF 1747 kb)

Supplementary Table 1

ASD and unaffected ASD sibling de novo variants. All 5,856 de novo variants in 3,982 individuals with ASD and 2,545 de novo variants in 2,078 unaffected siblings. Each row in the file represents one de novo variant in an individual. Descriptions of the column names are in the second tab. (XLSX 1915 kb)

Supplementary Table 2

ID/DD de novo variants. All 1,692 de novo variants in 1,284 individuals with ID/DD14-17. As with Supplementary Table 1, each row in the file represents one de novo variant in an individual. As noted in the DDD study, some de novo variants were observed in multiple unrelated individuals14. As with Supplementary Table 1, column name descriptions are in the second tab. (XLSX 398 kb)

Supplementary Table 6

Congenital heart disease and schizophrenia de novo variants. All 1,281 de novo variants in 362 individuals with congenital heart disease, and 640 de novo variants in 617 individuals with schizophrenia. As with Supplementary Tables 1 and 2, each row in the file represents one de novo variant in an individual. Likewise, descriptions of the column names are in the second tab. (XLSX 478 kb)

Supplementary Table 22

Gene summary statistics. Counts of class 1 de novo PTVs, transmitted and untransmitted singleton PTVs absent from ExAC, and singleton PTVs absent from ExAC from 404 cases and 3,654 controls grouped by gene. In total, there are 9,637 genes with at least one PTV from these categories. Descriptions of the column names are in the second tab. (XLSX 544 kb)

Supplementary Table 26

Transmitted and untransmitted PTVs from 4,319 ASD trios. All singleton, LofTee high-confidence PTVs absent from ExAC that were transmitted or untransmitted in 4,319 trios with ASD. Descriptions of the columns are found in the second tab. (XLSX 144 kb)

Supplementary Table 31

PTVs from Swedish case-control study. All singleton, LofTee high-confidence PTVs absent from ExAC that were present in 404 cases and 3,654 controls from Sweden. Descriptions of the columns are found in the second tab. (XLSX 88 kb)

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Kosmicki, J., Samocha, K., Howrigan, D. et al. Refining the role of de novo protein-truncating variants in neurodevelopmental disorders by using population reference samples. Nat Genet 49, 504–510 (2017).

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