Identification of ARHGAP35 Variants in ASD

This human study was approved by Institutional Review Boards at Children’s Wisconsin and Einstein Medical Center Philadelphia. Written informed consent including research analysis and photo publication if applicable was obtained for every participant. Exome sequencing was undertaken by Psomagen (Rockville, MD) and analyzed with VarSeq (Golden Helix, Bozeman, MT). In silico analysis of variants of interest included filtering for frequency <0.001 in the general population in gnomAD v2.1.1 [11 (link)] and for predicted effect upon the protein. The effect of missense variants on protein function was further analyzed by two combined analysis tools (CADD phred hg19 and REVEL). Samples were first analyzed for variants in known MAC and ASD genes as previously described [12 (link), 13 (link)]. Trio analysis in negative cases identified ARHGAP35 as a candidate in two families and screening for variants in this gene specifically identified one more case. Sanger sequencing was used to confirm variants and for segregation analysis. An additional case was identified through clinical genome sequencing and Matchmaker Exchange Databases [14 (link)]. Variants in ARHGAP35 were named based on reference sequence NM_004491.4 and human Genome Build hg19 and evaluated according to ACMG/AMP guidelines [15 (link)].