Identification of homozygous missense mutations in <i>ADD3</i> and <i>KAT2B</i> and effect of <i>ADD3</i> and <i>KAT2B</i> mutations on protein levels in fibroblasts.

<p>(A) Pedigree and segregation status of mutations found in <i>ADD3</i> and <i>KAT2B</i>. Discovery of <i>ADD3</i> mutations in family B and C was facilitated by GeneMatcher [<a href="" target="_blank">44</a>]. Half red coloured circles or squares denote patients with neurological defects and half blue coloured symbols denote patients with SRNS and cardiomyopathy. + symbols indicate non-mutated alleles. Mutations and segregation were confirmed by Sanger sequencing. (B) Exon structure of human <i>ADD3</i> cDNA (long isoform NP_058432) and domains of adducin-γ protein. The relative position of <i>ADD3</i> mutations to protein domains and exons are indicated (arrows). All mutations also affect the short isoform of <i>ADD3</i> (NP_001112). Below each mutation, the phylogenetic conservation of the altered amino acid residues is shown. (C) Exon structure of human <i>KAT2B</i> cDNA and domains of KAT2B protein. PCAF-HD, p300/CBP-associated factor homology domain; AT, acetyl transferase domain; B, Bromo domain. The relative position of <i>KAT2B</i> variation to protein domains and exons is indicated (arrow). The phylogenetic conservation of the altered amino acid residue is shown. (D, E) Adducin-γ (D) and KAT2B (E) protein levels in control and patient fibroblasts. Lysates of patient II-3 and II-6 (family A) fibroblasts and age-matched control fibroblasts (Ctrl 1 and 2) were analyzed by western blotting. Results were normalized to the loading control α-tubulin. Each quantification is shown in the lower panel (n = 3 independent experiments, student’s t-test).</p>