Supplementary Material for: Severe Progressive Autism Associated with Two de novo Changes: A 2.6-Mb 2q31.1 Deletion and a Balanced t(14;21)(q21.1;p11.2) Translocation with Long-Range Epigenetic Silencing of <i>LRFN5</i> Expression

In a 19-year-old severely autistic and mentally retarded girl, a balanced de novo t(14;21)(q21.1;p11.2) translocation was found in addition to a de novo 2.6-Mb 2q31.1 deletion containing 15 protein-encoding genes. To investigate if the translocation might contribute to developmental stagnation at the age of 2 years with later regression of skills, i.e. a more severe phenotype than expected from the 2q31.1 deletion, the epigenetic status and expression of genes proximal and distal to the 14q21.1 breakpoint were investigated in Ebstein Barr Virus-transformed lymphoblast and primary skin fibroblast cells. The 14q21.1 breakpoint was found to be located between a cluster of 7 genes 0.1 Mb upstream, starting with <i>FBXO33</i>, and the single and isolated <i>LRFN5</i> gene 2.1 Mb downstream. Only expression of <i>LRFN5</i> appeared to be affected by its novel genomic context. In patient fibroblasts, <i>LRFN5</i> expression was 10-fold reduced compared to <i>LRFN5</i> expressed in control fibroblasts. In addition, a relative increase in trimethylated histone H3 lysine 9 (H3K9M3)-associated DNA starting exactly at the translocation breakpoint and going 2.5 Mb beyond the <i>LRFN5</i> gene was found. At the <i>LRFN5</i> promoter, there was a distinct peak of trimethylated histone H3 lysine 27 (H3K27M3)-associated DNA in addition to a diminished trimethylated histone H3 lysine 4 (H3K4M3) level. We speculate that dysregulation of <i>LRFN5</i>, a postsynaptic density-associated gene, may contribute to the patient’s autism, even though 2 other patients with 14q13.2q21.3 deletions that included <i>LRFN5</i> were not autistic. More significantly, we have shown that translocations may influence gene expression more than 2 Mb away from the translocation breakpoint.