Identification of Germline Non-coding Deletions in XIAP Gene Causing XIAP Deficiency Reveals a Key Promoter Sequence

Abstract

X-linked inhibitor of apoptosis protein (XIAP) deficiency, also known as the X-linked lymphoproliferative syndrome of type 2 (XLP-2), is a rare immunodeficiency characterized by recurrent hemophagocytic lymphohistiocytosis, splenomegaly, and inflammatory bowel disease. Variants inXIAPincluding missense, non-sense, frameshift, and deletions of coding exons have been reported to cause XIAP deficiency. We studied three young boys with immunodeficiency displaying XLP-2-like clinical features. No genetic variation in the coding exons ofXIAPwas identified by whole-exome sequencing (WES), although the patients exhibited a complete loss of XIAP expression. Targeted next-generation sequencing (NGS) of the entire locus ofXIAPwas performed on DNA samples from the three patients. Molecular investigations were assessed by gene reporter expression assays in HEK cells and CRISPR-Cas9 genome editing in primary T cells. NGS ofXIAPidentified three distinct non-coding deletions in the patients that were predicted to be driven by repetitive DNA sequences. These deletions share a common region of 839bp that encompassed the first non-coding exon ofXIAPand contained regulatory elements and marks specific of an active promoter. Moreover, we showed that among the 839bp, the exon was transcriptionally active. Finally, deletion of the exon by CRISPR-Cas9 in primary cells reduced XIAP protein expression. These results identify a key promoter sequence contained in the first non-coding exon ofXIAP. Importantly, this study highlights that sequencing of the non-coding exons that are not currently captured by WES should be considered in the genetic diagnosis when no variation is found in coding exons.