A Functional Alternative Splicing Mutation in AIRE Gene Causes Autoimmune Polyendocrine Syndrome Type 1

Abstract

Autoimmune polyendocrine syndrome type 1 (APS-1) is a rare autosomal recessive disease defined by the presence of two of the three conditions: mucocutaneous candidiasis, hypoparathyroidism, and Addison’s disease. Loss-of-function mutations of the autoimmune regulator (AIRE) gene have been linked to APS-1. Here we report mutational analysis and functional characterization of an AIRE mutation in a consanguineous Chinese family with APS-1. All exons of the AIRE gene and adjacent exon-intron sequences were amplified by PCR and subsequently sequenced. We identified a homozygous missense AIRE mutation c.463G>A (p.Gly155Ser) in two siblings with different clinical features of APS-1. In silico splice-site prediction and minigene analysis were carried out to study the potential pathological consequence. Minigene splicing analysis and subsequent cDNA sequencing revealed that the AIRE mutation potentially compromised the recognition of the splice donor of intron 3, causing alternative pre-mRNA splicing by intron 3 retention. Furthermore, the aberrant AIRE transcript was identified in a heterozygous carrier of the c.463G>A mutation. The aberrant intron 3-retaining transcript generated a truncated protein (p.G155fsX203) containing the first 154 AIRE amino acids and followed by 48 aberrant amino acids. Therefore, our study represents the first functional characterization of the alternatively spliced AIRE mutation that may explain the pathogenetic role in APS-1.

Cohen’s h for detection of disease association with rare genetic variants

“…Another SNP, ss107794716 residing within the AIRE gene, was identified by both of RD and Cohen’s h but not OR (RD = 0.006, p = 0.034; Cohen’s h = 0.106, p = 0.019; OR = 7.04, p = 0.068). The association of this region and the AIRE gene with T1D was documented [ 35 – 37 ]. In addition, the magnitudes of p-values of significant rare variants obtained from Cohen’s h are the smallest.…”

Splicing analysis of rare/novel synonymous or intronic variants identified in ABCB11 heterozygotes presenting as progressive intrahepatic cholestasis with low γ‐glutamyltransferase

“…Primers are summarized in Table S2. A modified pcDNA3 minigene was used as the empty native minigene (vector), and was linearized by using Xba I and Xho I (Takara, Dalian, China). The wild‐type (WT) and mutant minigene were constructed by ligating the PCR products into linearized vector (Fig.…”

“…Minigene splicing assay is more frequently used . Cloning three consecutive exons, the affected exon and two flanking exons, into the minigene system may better mimic the procedure of pre‐RNA splicing in vivo , because it can retain the original ss sequences of upstream and downstream introns of the affected exon . The effects on pre‐RNA splicing can also be predicted by in silico tools, including MaxEntScan (MES), Splice‐Site Prediction by Neural Network (NNSplice), Human Splicing Finder (HSF), Splice‐Port, Exonic Splicing Enhancers (ESE) Finder, and RESCUE‐ESE, but they are insufficient.…”

AAV9‐mediated AIRE gene delivery clears circulating antibodies and tissue T‐cell infiltration in a mouse model of autoimmune polyglandular syndrome type‐1

“…11 To date, more than 100 APS-1 causing mutations have been identified which vary from substitutions, insertions and deletions to splice-site mutations. [12][13][14] Several studies have demonstrated that AIRE/ Aire is expressed mainly by the thymus, in a subpopulation of medullary thymic epithelial cells (mTECs), AIRE + mTECs. 9,[15][16][17] AIRE/Aire promotes self-tolerance in the thymus by regulating the promiscuous expression of a wide array of tissuespecific antigens (TSAs).…”