High allele frequency of the p.Q258X mutation and identification of a novel mis-splicing mutation in the STAR gene in Korean patients with congenital lipoid adrenal hyperplasia

Abstract

Objective: Steroidogenic acute regulatory (STAR) protein plays a crucial role in steroidogenesis, and mutations in the STAR gene cause congenital lipoid adrenal hyperplasia (CLAH). This study investigated the STAR mutation spectrum and functionally analyzed a novel STAR mutation in Korean patients with CLAH. Methods: Mutation analysis of STAR was carried out in 25 unrelated Korean CLAH patients. A region of STAR comprising exons 4-7 was cloned from human genomic DNA into an expression vector, followed by site-directed mutagenesis and transient expression in COS7 cells. The splicing pattern was analyzed by in vitro transcription, and each transcript was functionally characterized by measuring pregnenolone production in COS7 cells cotransfected with the cholesterol side chain cleavage system. Results: Mutation p.Q258X was identified in 46 of 50 alleles (92%); mutation c.653COT was detected in two alleles (4%); and mutations p.R182H and c.745-6_810del were found in one allele (2%). Reverse transcriptase-PCR products amplified from a patient heterozygous for compound c.653COT and c.745-6_810del mutation revealed multiple alternatively spliced mRNAs. In vitro expression analysis of a minigene consisting of exons 4-7 containing the c.653COT yielded two transcripts in which exon 6 or exons 5 and 6 were skipped. The encoded proteins exhibited defective pregnenoloneproducing ability. The c.745-6_810del mutation led to full and partial intron retention. Conclusions: p.Q258X is the most common STAR mutation in Korea. A previously reported c.653COT variant was found to cause aberrant splicing at the mRNA level, resulting in perturbation of STAR function. The c.745-6_810del mutation also resulted in aberrant splicing.

p.R182C mutation in Korean twin with congenital lipoid adrenal hyperplasia

“…In patients with transition arginine at codon 182 to other amino acid reported late onset of symptoms and mild symptoms1,2,7), but some of them showed symptoms early7). Mutation of p.Q258X resulted in premature stop codon and loss of StAR activity with formation of truncated protein10). Although there was no report functional study of p. R182C, failing formation of the cholesterol biding pocket was described in p.R182H2,7).…”

Early steps in steroidogenesis: intracellular cholesterol trafficking

“…Many StAR mutations have been found in lipoid CAH patients, but about 65-70% of affected Japanese alleles and virtually all affected Korean alleles carry the mutation Q258X ( 200,201,220,246,(265)(266)(267). The Q258X carrier frequency in these countries is about 1 in 300 ( 201,266,267 ) so that 1 in every 250,000 to 300,000 newborns is affected, or about 500 patients in Japan and Korea. Other genetic clusters are found among Palestinian Arabs, carrying the mutation R182L ( 201 ), in eastern Saudi Arabia, carrying R182H ( 261 ), and in parts of Switzerland, carrying the mutation L260P ( 268 ).…”

ACTH Action on StAR Biology

“…The finding that STAR mutations are the genetic basis for lipoid CAH was key to establishing the essential role for StAR in ACTH-stimulated steroidogenesis as well as gonadotropin-stimulated steroidogenesis (Lin et al, 1995; Caron et al, 1997). A common mutation found in the STAR gene is a nonsense mutation, Q258X, which results in truncation of the last 28 amino acids (Nakae et al, 1997; Kim et al, 2011). Expression of C-terminal truncated forms of StAR in COS-1 cells confirmed that the loss of C-terminal helix, within the START domain, results in an inactive protein (Arakane et al, 1996; Wang et al, 1998).…”