Aneuploid rescue precedes X-chromosome inactivation and increases the incidence of its skewness by reducing the size of the embryonic progenitor cell pool

Abstract

STUDY QUESTION Do monosomy rescue (MR) and trisomy rescue (TR) in preimplantation human embryos affect other developmental processes, such as X-chromosome inactivation (XCI)? SUMMARY ANSWER Aneuploid rescue precedes XCI and increases the incidence of XCI skewness by reducing the size of the embryonic progenitor cell pools. WHAT IS KNOWN ALREADY More than half of preimplantation human embryos harbor aneuploid cells, some of which can be spontaneously corrected through MR or TR. XCI in females is an indispensable process, which is predicted to start at the early-blastocyst phase. STUDY DESIGN, SIZE, DURATION We examined the frequency of XCI skewness in young females who carried full uniparental disomy (UPD) resulting from MR or TR/gamete complementation (GC). The results were statistically analyzed using a theoretical model in which XCI involves various numbers of embryonic progenitor cells. PARTICIPANTS/MATERIALS, SETTING, METHODS We studied 39 children and young adults ascertained by imprinting disorders. XCI ratios were determined by DNA methylation analysis of a polymorphic locus in the androgen receptor gene. We used Bayesian approach to assess the probability of the occurrence of extreme XCI skewness in the MR and TR/GC groups using a theoretical model of 1–12 cell pools. MAIN RESULTS AND THE ROLE OF CHANCE A total of 12 of 39 individuals (31%) showed skewed XCI. Extreme skewness was observed in 3 of 15 MR cases (20%) and 1 of 24 TR/GC cases (4.2%). Statistical analysis indicated that XCI in the MR group was likely to have occurred when the blastocyst contained three or four euploid embryonic progenitor cells. The estimated size of the embryonic progenitor cell pools was approximately one-third or one-fourth of the predicted size of normal embryos. The TR/GC group likely had a larger pool size at the onset of XCI, although the results remained inconclusive. LIMITATIONS, REASONS FOR CAUTION This is an observational study and needs to be validated by experimental analyses. WIDER IMPLICATIONS OF THE FINDINGS This study provides evidence that the onset of XCI is determined by an intrinsic clock, irrespectively of the number of embryonic progenitor cells. Our findings can also be applied to individuals without UPD or imprinting disorders. This study provides a clue to understand chromosomal and cellular dynamics in the first few days of human development, their effects on XCI skewing and the possible implications for the expression of X-linked diseases in females. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by the Grants-in-aid for Scientific Research on Innovative Areas (17H06428) and for Scientific Research (B) (17H03616) from Japan Society for the Promotion of Science (JSPS), and grants from Japan Agency for Medical Research and Development (AMED) (18ek0109266h0002 and 18ek0109278h0002), National Center for Child Health and Development and Takeda Science Foundation. The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER Not applicable.

Random X chromosome inactivation in patients with Klinefelter syndrome

“…We demonstrated that the XCI ratios of KS patients followed a normal distribution. Furthermore, the frequencies of skewed XCI and extremely skewed XCI in KS patients were comparable to previous data obtained from healthy young women [7]. These findings are consistent with previous reports by Zitzmann et al [15] and Zinn et al [16], which documented apparently random XCI in KS patients.…”

“…The XCI ratios of the two cases were 13.3%:86.7% and 91.6%:8.4%. The frequencies of skewed XCI and extremely skewed XCI in the KS cohort were comparable to those in healthy young women [7] (11.1% vs. 11.5%, p = 0.641 and 5.6% vs. 1.9%, p = 0.342, respectively).…”

“…The methods were as described previously [18]. The accuracy of the experiments in our laboratory has previously been confirmed by analyzing more than 200 DNA samples from healthy individuals [7]. The XCI ratio was expressed as the ratio of inactive X chromosomes with less CAG repeats to those with more repeats [14].…”

“…The distribution of XCI ratios of KS patients was statistically analyzed by employing the Shapiro-Wilk test. In addition, the frequency of skewed XCI (≥ 80%:20% or ≤ 20%:80%) and extremely skewed XCI (≥ 90%:10% or ≤ 10%:90%) in the KS cohort was compared with our previous data obtained from 208 healthy young women [7], by using Fisher's exact test. Statistical analyses were performed with the IBM SPSS Statistics software (version 24.0).…”

“…These results indicated that XCI starts when a blastocyst contains 10-12 embryonic progenitor cells. Recently, we showed that the frequency of skewed XCI is increased in individuals with uniparental disomy (UPD) due to monosomy rescue or trisomy rescue/gamete complementation [7]. We proposed that life-saving aneuploid rescue increases the frequency of skewed XCI by reducing the size of embryonic progenitor cell pools at the onset of XCI.…”