The impact of hypothalamic overexpression of Makorin ring finger protein 3 (MKRN3) in postpubertal mice

Date
2022
DOI
Authors
McKnight, Kali
Version
OA Version
Citation
Abstract
BACKGROUND: The onset of puberty is marked by the activation of the hypothalamic-pituitary-gonadal (HPG) axis. Many neuroendocrine factors affect the timing of puberty and maintenance of reproduction, including genetic factors. Loss-of-function mutations in Makorin Ring Finger Protein 3 (MKRN3) is an important cause of central precocious puberty. MKRN3 declines prior to the onset of puberty and serves as an inhibitor to the onset of puberty. However, its potential role in delayed puberty has not been defined. AIMS AND METHODS: The aim of this study was to observe the age-specific effects of Mkrn3 overexpression on the reproductive phenotype of male and female mice. It was hypothesized that overexpression of Mkrn3 in the arcuate nucleus of mice would inhibit the HPG axis and lead to central hypogonadism in female mice, but not male mice, because of its possible influence on neuropeptides pertinent to the maintenance of reproduction. A model of Mkrn3 overexpression was generated using a recombinant adeno-associated virus (rAAV-Mkrn3) that was stereotaxically injected bilaterally, targeting the arcuate nucleus of wild-type C57/B6 postpubertal male and female mice. A control group of mice was also injected with the virus (rAAV-EGFP), in which EGFP is enhanced green fluorescent protein. Initially, the estrous cyclicity was monitored in gonad-intact female mice. Stimulation testing with peripherally administered senktide, kisspeptin, and/or GnRH was then performed on gonad-intact female mice, ovariectomy plus estradiol-replaced (OVX+E2) female mice, and gonad-intact male mice. Serum luteinizing hormone (LH) levels were assessed at baseline and at 20, 40, and 60 minutes following neuropeptide administration. Finally, whole brains were collected for protein expression for Mkrn3 and EGFP by immunohistochemistry. RESULTS: The rAAV-Mkrn3 female mice spent a greater percentage of time in diestrus compared with the rAAV-EGFP female mice. Gonad-intact and OVX+E2 rAAV-Mkrn3 female mice demonstrated a statistically significant decrease in peak serum LH levels in response to the neurokinin 3 receptor (NK3R) agonist, senktide, compared with female mice stereotaxically injected with rAAV-EGFP. Serum LH levels in response to peripheral administration of kisspeptin and gonadotropin-releasing hormone (GnRH) were not different between rAAV-Mkrn3 and rAAV-EGFP female mice. Gonad-intact male mice demonstrated statistically similar LH levels upon administration of senktide, kisspeptin, and GnRH for both rAAV-Mkrn3 and rAAV-EGFP mice. CONCLUSIONS: This study aimed to understand the implications of MKRN3 overexpression on the HPG axis after pubertal onset and to determine if there were specific developmental windows by which MKRN3 could exert an effect on the reproductive axis. The findings of this study indicate that hypothalamic Mkrn3 overexpression may be directly or indirectly impacting the neuropeptides of the kisspeptin, neurokinin B, and dynorphin (KNDy) neurons of the arcuate nucleus, especially NK3R and/or kisspeptin. This effect could be due to the E3 ubiquitin ligase activity and/or the RNA-binding activity of MKRN3. Because hypothalamic Mkrn3 overexpression partially suppressed the HPG axis in postpubertal female mice, MKRN3 could be a yet unrecognized cause of hypogonadotropic hypogonadism (HH) in humans. This suggests that MKRN3 may have potential clinical implications such as serving as a novel therapeutic agent for children with central precocious puberty (CPP) and other disorders in which the inhibition of the HPG axis is desired.
Description
License