FMR1 is a gene on the X chromosome that contains an expansion-prone CGG?CCG repeat in its 5'untranslated region. Alleles with 55-200 repeats are considered to be Fragile X premutation (PM) alleles. The frequency of such alleles in women ranges from 1 in 113 to 1 in 250. Carriers of such alleles are at risk for Fragile X Primary Ovarian Insufficiency (FXPOI) that is seen in up to 28% of women who carry such alleles. FXPOI accounts for ~11.5% of familial cases of infertility and 3.5% of idiopathic cases. Even without a diagnosis of FXPOI, the average age at menopause of all women with the premutation is ~5 yrs earlier than their siblings without the PM. Thus, female PM carriers not only have increased fertility problems, but are at greater risk of cardiovascular disease, Alzheimer disease, osteoporosis and other problems that are seen at higher frequency in menopausal women. Hormonal assessment of PM carriers indicates that FSH is elevated, inhibins and anti-Mullerian hormone (both made by granulosa cells) are reduced. Preliminary data from a knock-in Fragile X PM mouse model containing ~130 CGG repeats suggest a number of ovarian problems. These include early losses of immature follicles and reductions in corpora lutea. Advanced follicles were smaller in PM mice and showed a higher rate of atresia. This was associated with a smaller than normal number of granulosa cells (GCs). In addition, PM oocytes showed abnormal nuclear morphology, reduced levels of gap junction proteins, abnormal staining of their zona pellucida, a primarily nuclear location of the normally cytoplasmic fragile X mental retardation protein (FMRP) and high levels of ubiquitinated proteins that often accumulated in the nucleus or perinuclear region. A high incidence of large cysts was also seen in PM mice. Thus our mice exhibit signs consistent with ovarian insufficiency and have unusual ovarian changes that could contribute to this ovarian dysfunction. These mice may therefore provide a good model of FXPOI.
Aim 1 uses transplantation approaches to address whether the PM in the ovary alone is sufficient to produce the FXPOI-like and aberrant ovarian features or instead requires the PM in the hypothalamic/pituitary unit.
Aim 2 focuses on the abnormal FMRP, ubiquitin, and gap junctions as well as the abnormalities in zona pellucida composition to attempt to uncover the mechanism(s) responsible for the follicle decline.
FMR1 premutation (PM) alleles have 55-200 repeats in the 5'UTR of the gene;~1:130 women have such alleles. These women are at risk of premature primary ovarian insufficiency (FXPOI). Changes in cycling PM carriers show elevated FSH coupled with reduced inhibins, and anti-Mullerian hormone levels, all of which are thought to signify a lower ovarian follicle reserve. However, very little is known as to how ovarian problems arise from the mutated FMR1 gene, or even which ovarian cells are initiating the cascade to ovarian decline. This proposal aims to fill those gaps using Fragile X PM knock-in mice with ~130 CGG repeats in their Fmr1 gene. Preliminary results suggest that the PM ovary is consistent with FXPOI, exhibiting premature follicle loss that could stem from problems in the production of follicular somatic cells (granulosa cells) and/or their communication with the oocyte. The proposed studies will begin to unravel the mechanisms by which PM results in compromised ovarian function and reduced ovarian lifespan.