To evaluate further the genes with multiple truncating variants, we examined segregation of the variants in the families in which they were observed. Some of these results have been previously published16-21. In brief, truncating variants in AP1S2, CUL4B, BRWD3, UPF3B, ZDHHC9 and SLC9A6 segregated completely with mental retardation in the families in which they were identified; that is, each truncating variant was present in all genotyped subjects with mental retardation and absent in unaffected males (Supplementary Fig. 1). We sequenced all the coding exons of these six genes in control X chromosomes and did not find the truncating variants detected in XLMR-affected subjects or any other truncating variants (Table 3). The clustering of multiple different truncating variants in these genes, the evidence for segregation with mental retardation and the absence of truncating variants in controls indicate strongly that AP1S2, CUL4B, BRWD3, UPF3B, ZDHHC9 and SLC9A6 are XLMR genes. Five missense or in-frame variants in CUL4B, ZDHHC9 and SLC9A6 also showed evidence of involvement in mental retardation17,19,21. Mental retardation–causing variants in AP1S2, CUL4B, BRWD3, UPF3B, ZDHHC9 and SLC9A6 together account for the disease in 22 (10.6%) families out of the 208 screened.
