Endocrine Abstracts

Most mutations found in the androgen receptor (AR) are associated with some form of androgen insensitivity syndrome (AIS), and correlate with a loss of either hormone, DNA or coactivator binding. For some mutations in the ligand binding domain (LBD) of the AR, it is unclear what the underlying mechanism of the AIS could be. In the beginning of 2017, a new dimerization interface was uncovered in a crystal structure of the ligand-binding domain (LBD) which provides a possible mechanism for several unexplained cases of AIS.¹ We now want to uncover the physiological relevance of this newly uncovered dimerization method as well as its role in AR functioning. We generated a mouse model based on a W752R mutation located in the LBD dimerization interface, described in two siblings with androgen insensitivity. In vitro, AR W752R is well expressed, binds androgens with high affinity and has proven an impaired LBD dimerization potential. Although at lower androgen concentrations AR W752R is less potent on androgen reporter genes, at higher concentrations it has strong transactivating potential similar to wild type AR. Via the CRISPR/Cas9 technique, we introduced the W to R mutation into C57BL/6J mice. This model is called AR Lmon (for monomeric mutation in the LBD). To exclude effects caused by off target mutations that may be incorporated during the CRISPR/Cas method, we performed backcrossing of the mice until at least the third generation. The male AR Lmon mice show an external female phenotype with female ano-genital distance and nipple development. Testes are present, smaller than wild type, but not as small as seen in our complete androgen knockout mice (ARKO). Analogous to complete AIS (CAIS), the Lmon mice did not developed epididymis, seminal vesicle or musculus levator ani. These observations suggest that the interruption of the dimerization of the AR via the LBD leads to a severe androgen insensitivity phenotype. The growth curve of the Lmon mice is comparable to the wild type females and lower than the one of normal males. However, surprisingly, these mice start to gain weight after the age of 12 weeks. Dissection at 16 weeks showed an increase in subcutaneous, retroperitoneal and gonadal fat comparing to wild type and ARKO males. Preliminary measurement of testosterone and LH concentrations in circulation of two animals uncovered that the production of both is highly elevated in Lmon mice, indicating a disrupted hypothalamus-pituitary-gonadal axis similar to what is observed in CAIS patients. In conclusion, a mouse model of the W752R mutation phenocopies the CAIS, despite the fact that in vitro the AR still can transactivate reporter genes. To uncover the exact place of LBD dimerization in the normal AR functioning and male development, further investigations are necessary.

Reference: 1. Nadal, M. et al. Structure of the homodimeric androgen receptor ligand-binding domain. Nat. comm. 8, 14388 (2017).