Paths were partitioned by family member. Under the assumption that SNV and indel variants occur randomly across coding regions, larger genes will be more

likely to accumulate higher numbers of such variants. In addition, (1) the proportion of a gene that is included in the design of the capture reagents and (2) nonuniform capture coverage across the target will influence the expected numbers of variants of a gene or group of genes. To address these issues, we based our expectation of number of variants per gene (or group of genes) on the distribution of observed rare synonymous mutations in the 686 parents. Specifically, 7,051 of the 63,080 (or 11.18%) of all rare synonymous SNVs fell within the FMRP-associated genes (Table 6). We set 11.18% as the expected proportion for Venetoclax LGD variants in FMRP-associated genes and used a binomial test to assign p values to the observed overlap between FMRP-associated genes and LGD variants in probands and their unaffected siblings and assigned p values for INCB018424 order the overlap with missense variants similarly (Table 5). CNV candidate genes (Gilman et al., 2011) were obtained through a greedy optimization procedure that selected the most interconnected (in the context of a whole genome molecular network) subset of genes from the set of genes affected by a de novo deletion or duplication

in autistic probands. The molecular network utilized cumulative expert and experimental knowledge that was heavily biased toward what had been studied others such that it was difficult to accurately quantify. To measure the significance of the observed overlap between the 72 CNV candidates and the FMRP-associated genes, we performed a permutation test: random CNV regions were selected, preserving the number of genes as in the real CNVs, the 72 most interconnected genes were identified using the greedy optimization (allowing at most 2 genes per CNV region), and the overlap with FMRP-associated genes was recorded (Gilman et al.,

2011). We repeated this procedure 10,000 times and built an empirical distribution for the number of FMRP-associated genes if the CNVs were taken as random. Only 4 of 10,000 permutations produced an overlap equal to or larger than the observed 13 FMRP-associated genes. This work was supported by grants from the Simons Foundation (SF51 and SF235988) to M.W. and by a grant from the NIH (5RC2MH090028-02) to M.W. and W.R.M. We are grateful to all of the families at the participating SFARI Simplex Collection (SSC) sites, as well as the principal investigators (A. Beaudet, R. Bernier, J. Constantino, E. Cook, E. Fombonne, D. Geschwind, D. Grice, A. Klin, R. Kochel, D. Ledbetter, C. Lord, C. Martin, D. Martin, R. Maxim, J. Miles, O. Ousley, B. Pelphrey, B. Peterson, J. Piggot, C. Saulnier, M. State, W. Stone, J. Sutcliffe, C. Walsh, and E. Wijsman).