Promoter-Dependent Activity on Androgen Receptor N-Terminal Domain Mutations in Androgen Insensitivity Syndrome

Abstract

Androgen receptor (AR) mutations are associated with androgen insensitivity syndrome (AIS). Missense mutations identified in the AR-N-terminal domain (AR-NTD) are rare, and clinical phenotypes are typically mild. We investigated 7 missense mutations and 2 insertion/deletions located in the AR-NTD. This study aimed to elucidate the pathogenic role of AR-NTD mutants in AIS and to use this knowledge to further define AR-NTD function. AR-NTD mutations (Q120E, A159T, G216R, N235K, G248V, L272F, and P380R) were introduced into AR-expression plasmids. Stably expressing cell lines were established for del57L and ins58L. Transactivation was measured using luciferase reporter constructs under the control of GRE and Pem promoters. Intrinsic fluorescence spectroscopy and partial proteolysis studies were performed for mutations which showed reduced activities by using a purified AR-AF1 protein. Pem-luciferase reporter activation was reduced for A159T, N235K, and G248V but not the GRE-luciferase reporter. Protein structure analysis detected no significant change in the AR-AF1 region for these mutations. Reduced cellular expression and transactivation activity were observed for ins58L. The mutations Q120E, G216R, L272F, P380R, and del57L showed small or no detectable changes in function. Thus, clinical and experimental analyses have identified novel AR-signalling defects associated with mutations in the structurally disordered AR-NTD domain in patients with AIS.

Predicting puberty in partial androgen insensitivity syndrome: Use of clinical and functional androgen receptor indices

“…While the additional use of yeast two-hybrid assays provides mechanistic insights into the nature of the functional defect in AR signalling, it is evident that the use of in vitro assays of androgen action used in this study is not sufficiently informative to predict how puberty will develop in PAIS associated with a particular AR mutation. This may be due to variable responses according to different promoters used in reporter gene assays under the conditions employed [ 21 , 22 ]. The complexity of nuclear receptor coregulator dynamics is now well established, where the activity of the receptor itself is influenced by the DNA elements bound, together with coregulator expression and recruitment.…”

Comparing the rules of engagement of androgen and glucocorticoid receptors

“…Physiological evidence for allosteric effects in the AR comes from the functional analysis of AR-NTD mutations, and detected in androgen insensitivity patients that affect the functioning of AR in an ARE-specific way [62]. In addition, the deletion of the (23)FQNLF(27) motif which is involved in NTD-LBD communications in the AR has an effect on transactivation via classical AREs, but has little effect on reporters based on selAREs [63].…”

Clinical and molecular characteristics in 15 patients with androgen receptor gene mutations from South China

“…The c.171_172insCTG insertion (ins58L) is a reported mutation observed in exon 1 (Ferlin et al., ; Tadokoro‐Cuccaro et al., ). This leucine insertion mutation was reported to be associated with male infertility and various degrees of hypospadias.…”