TP63 gain-of-function mutations cause premature ovarian insufficiency by inducing oocyte apoptosis
Chengzi Huang, … , Zi-Jiang Chen, Shidou Zhao
Chengzi Huang, … , Zi-Jiang Chen, Shidou Zhao
Published March 1, 2023
Citation Information: J Clin Invest. 2023;133(5):e162315. https://doi.org/10.1172/JCI162315.
View: Text | PDF
Research Article Genetics Reproductive biology

TP63 gain-of-function mutations cause premature ovarian insufficiency by inducing oocyte apoptosis

Abstract

The transcription factor p63 guards genome integrity in the female germline, and its mutations have been reported in patients with premature ovarian insufficiency (POI). However, the precise contribution of the TP63 gene to the pathogenesis of POI needs to be further determined. Here, in 1,030 Chinese patients with POI, we identified 6 heterozygous mutations of the TP63 gene that impaired the C-terminal transactivation-inhibitory domain (TID) of the TAp63α protein and resulted in tetramer formation and constitutive activation of the mutant proteins. The mutant proteins induced cell apoptosis by increasing the expression of apoptosis-inducing factors in vitro. We next introduced a premature stop codon and selectively deleted the TID of TAp63α in mice and observed rapid depletion of the p63+/ΔTID mouse oocytes through apoptosis after birth. Finally, to further verify the pathogenicity of the mutation p.R647C in the TID that was present in 3 patients, we generated p63+/R647C mice and also found accelerated oocyte loss, but to a lesser degree than in the p63+/ΔTID mice. Together, these findings show that TID-related variants causing constitutive activation of TAp63α lead to POI by inducing oocyte apoptosis, which will facilitate the genetic diagnosis of POI in patients and provide a potential therapeutic target for extending female fertility.

Authors

Chengzi Huang, Simin Zhao, Yajuan Yang, Ting Guo, Hanni Ke, Xin Mi, Yingying Qin, Zi-Jiang Chen, Shidou Zhao

×

Figure 7

Evaluation of fertility and oocyte quality in p63+/R647C mice.

Options: View larger image (or click on image) Download as PowerPoint
Evaluation of fertility and oocyte quality in p63+/R647C mice. (A) Cumul...
(A) Cumulative number of pups obtained from WT females (n = 5) and p63+/R647C females (n = 5) crossed with WT males for a period of 3 months. (B and C) The numbers of pups per litter and the number of litters per mouse were recorded for each group (n = 5) in the fertility test. (D) Number of superovulated oocytes per mouse obtained from the 2 groups (n = 5 for each genotype). (E) Morphology of superovulated oocytes obtained from 3-week-old WT and p63+/R647C mice. Scale bars: 100 μm. (F) Percentages of MII oocytes with PB1 emission in WT and p63+/R647C oocytes. Three mice for each genotype were used for each independent experiment, and 3 independent experiments were conducted. (G) The morphology of spindle and chromosome organization in WT and p63+/R647C oocytes. Anti–α-tubulin antibody (green) was used to stain the spindles. Chromosomes were counterstained with DAPI (blue). Scale bars: 20 μm. (H) Percentages of oocytes with spindle/chromosome defects in WT and p63+/R647C oocytes. Three mice of each genotype were used for each independent experiment, and 3 independent experiments were performed. (I) Oocyte MMP shown by JC-1 staining in the 2 groups. Red fluorescence indicates JC-1 aggregates with higher MMP, and green fluorescence indicates JC-1 monomers with lower MMP. Scale bars: 20 μm. (J) Quantification of the ratio of red to green fluorescence intensity in the 2 groups. Three mice for each genotype were used for each independent experiment, and 3 independent experiments were performed. (A–D, F, H, and J) Data are presented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, by unpaired, 2-tailed Student’s t test.