Suzanne de Bruijn

89 Homozygous KIAA1549 variants are associated with retinitis pigmentosa Assessing the function of KIAA1549 To investigate whether KIAA1549 plays a role in ciliogenesis, an in vitro study was performed in which KIAA1549 was knocked down in hTERT-RPE1 cells with two different siRNAs targeting the long transcript.Theefficiencyof KIAA1549 knockdownwas validated by qPCR analysis, and induced ciliogenesis was studied using immunocytochemistry. The percentage of ciliated cells and average cilium length showed no significant difference when comparing cells treated with KIAA1549- targeting siRNAs or the non- targeting siRNA ( Figure S1 ), suggesting that KIAA1549 does not have a function in the formation of cilia however KIAA1549 may be involved in other processes that are performed at the primary cilium. DISCUSSION In this study, we report on two families in which autosomal recessive RP is associated with homozygous variants in KIAA1549 . The retinal phenotype in both families is typical for RP, and patients’ complaints started with night blindness with subsequent constriction of the visual field. Fundus examination revealed the hallmark RP features. However, patients in Family A are more severely affected compared to the patient in Family B, which is displayed in a lower age at onset, severely constricted visual fields, and more severely reduced electroretinography (ERG) responses. In Family A, a homozygous frameshift variant was found in exon 1 (c.52del; p.(Arg18Alafs*64)) by WES. Although putative alternative start codons are present in exon 2, exon 1 encodes the signal peptide of the protein (aa 1-60). Therefore, a shorter protein is potentiallymislocalized, impairingprotein function. In Family B, a homozygous missense variant was found in exon 14 (c.4686C>A; p.(His1562Gln)), that affects a highly conserved region of the protein. The damaging nature of these variants is supported by a probability of loss of function intolerance (pLI) score of 1.00 (Scale 0-1) in gnomAD (accessed on 1 st June 2018) and that no homozygous variants have been reported in the entire KIAA1549 gene. Combined, this suggests that the identified KIAA1549 variants in both families can be associated to the RP-phenotype of the patients. Regardless, the presence of pathogenic variants present in non-coding regions uncovered by WES cannot be ruled out. KIAA1549 encodes a transmembrane protein and is described to be predominantly expressed in the brain and is involved in oncogenesis when fused to BRAF . 5,23 BRAF- KIAA1549 in-frame fusion genes are caused by a 2 Mb tandem duplication at 7q34, and are found to induce BRAF kinase activity and consequently, activation of the