br Concluding remarks br Conflict of

Concluding remarks
Conflict of interest
Acknowledgment This work was supported by the H.E.B. Endowed Chair in Marine Science (to P.T.) and University of Texas Marine Science Institute endowment-supported scholarships (to A.C).
Introduction Disorders of sex development (DSD) are defined as congenital conditions in which development of chromosomal, gonadal or anatomical sex is atypical. These disorders are classified into three major categories: sex chromosome DSD, 46,XX DSD and 46,XY DSD [1]. The 46,XY DSD are characterized by ambiguous or female external genitalia, caused by incomplete intrauterine masculinization, and the presence or absence of Mullerian structures [2]. 46,XY DSD can result from impaired production of testosterone, decreased conversion of testosterone to dihydrotestosterone or impaired peripheral action of these hormones [3]. Androgen insensitivity syndrome (AIS; OMIM#300068), the most common known cause of 46,XY DSD is defined as a condition resulting from complete or partial resistance to the biological actions of androgens in an XY man or boy with normal testis determination and production of age-appropriate androgen concentrations [4]. The clinical of manifestations AIS could range from phenotypic females (complete form; CAIS) to milder degrees of undervirilization (partial form; PAIS) or the mild form (MAIS) in which males have gynecomastia and/or infertility [5]. CAIS, which has been previously termed testicular feminization syndrome, was first described by Morris in 1953 [6]. The characteristic features of CAIS are normal female phenotype, normal breast development, absent or sparse pubic and axillary hair, an absence of the uterus and ovaries, and a short blind-ending vagina. The estimated prevalence of CAIS ranges from 1:20,400 to 1:99,100 genetic males on the basis of proven molecular diagnosis [7]. CAIS is an X-linked recessive genetic disease that is a result of mutations in the Androgen Receptor gene (AR; OMIM#313700). The AR gene is located on chromosome Xq11-12 and consists of 8 exons, encoding a protein of 920 protein phosphatase inhibitor [8]. The AR protein is an intracellular transcription factor that is a member of the nuclear receptor superfamily. Like all nuclear receptors, the AR protein consists of four major functional domains: a N-terminal domain (NTD) encoded by exon 1, a DNA-binding domain (DBD) encoded by exons 2 and 3, a hinge region encoded by exons 3 and 4, and a ligand-binding domain (LBD) encoded by exons 4–8 [9]. To date, more than 300 different mutations have been identified in the AR gene (www.mcgill/androgendb/). The most common molecular defects are missense mutations leading to amino acid substitutions in the AR protein. However, splice site mutations and complete deletions, as well as small insertion and deletions leading to premature termination of the protein are also reported [10]. In the present study, we studied one Chinese family with two siblings affected with complete androgen insensitivity syndrome and identified a splice acceptor site mutation (c.1769-1G > C) in AR gene from patients. Moreover, the consequence of this splice site mutation was investigated by computational analysis and functional study.
Materials and methods
Results
Discussion The production of mature RNA in eukaryotes requires an accurate removal of intervening sequences by splicing. Splicing of human pre-mRNAs is mediated by conserved but highly degenerate signal sequences at the splice sites. These signals include the MAG/GTRAGT consensus (M is A or C, R is purine, backslash indicates the exon–intron boundary) at the splice donor site, and the YAG/R consensus (Y is pyrimidine) at the splice acceptor site, with upstream polypyrimidine tract (PPT) and the branch point sequence (BPS; mammalian consensus YNCTRAY) [13]. The splicing signals of acceptor sites are recognized by RNA–protein interactions involving splicing factor 1 (SF1) and U2 small nuclear RNP auxiliary factor (U2AF) [14]. U2AF is a heterodimer, comprising of a 65-kDa and a 35-kDa subunit [15].U2AF65 binds the PPT immediately downstream of the BPS, while U2AF35 interacts with the AG dinucleotides in a sequence-specific fashion and can stabilize U2AF65 binding to pre-mRNAs [16], [17]. The most common outcome of mutations affecting splice sites is exon skipping, followed by cryptic splice site activation [18].