PCR amplification of exon 3 of APOPT1 was unsuccessful using genomic DNA from individual S4 (Figure S4A), suggesting a homozygous deletion of the corresponding genomic region, and no mutation was identified in other exons. Since we successfully generated PCR products of exons 2 and 4, we assume that the deletion does not extend beyond 15,328 bp, corresponding to the distance between oligonucleotide primers 2R and 4F. Accordingly, analysis of the cDNA retrotranscribed from the mutant transcript showed the absence of the mRNA portion encoded by exon 3 (Figures S4B and S4C). The deletion of exon 3 causes a change in the reading frame of APOPT1 and is predicted to result in the introduction of a premature stop codon (p.Glu121Valfs∗6). In individual S5, we found a homozygous c.353T>C mutation transition, predicting a p.Phe118Ser substitution. Phe118 is highly conserved, with mutation to a serine residue being predicted as extremely deleterious by several bioinformatics tools (Figure S5). In individual S6, we identified two heterozygous mutations: the same c.235C>T change present in individuals S1 and S2 and a three-nucleotide deletion (c.370_372delGAA) causing the elimination of a highly conserved amino acid residue (p.Glu124del). Parents were shown to be heterozygous carriers of one mutation, and a healthy sibling was heterozygous for the nonsense mutation. Details of the APOPT1 mutations and corresponding changes in the protein are summarized in Figure 3A and Table 3.