Timing of the absence of FMR1 expression in full mutation chorionic villi

Abstract.

Fragile X syndrome is caused by the expansion of the CGG repeat in the 5' untranslated region of the FMR1 gene. This expansion leads to methylation of the FMR1 promoter region thereby blocking FMR1 protein (FMRP) expression. Prenatal diagnosis can be performed on chorionic villi samples (CVS) by Southern blot analysis. Alternatively, for males, an immunohistochemical method has been introduced for CVS. In this study, we have used this immunocytochemical method for CVS in full mutation male fetuses at different times of gestational age, varying from 10.0–12.5 weeks, and in two cases of full mutation female fetuses (>13 weeks). FMRP expression studies in CVS from full mutation male fetuses (10.0–12.5 weeks) illustrate the timing of the disappearance of FMRP expression in these CVS. Until approximately 10 weeks of gestation, FMRP is expressed normally in full mutation male CVS, whereas FMRP is completely absent at 12.5 weeks of gestation. FMRP expression in full mutation female CVS (>13 weeks) is completely absent in a number of villi, whereas other villi show normal FMRP expression. Unlabelled villi can only be present in the absence of the expression of the full mutation FMR1 gene on one X-chromosome together with the X-inactivation of the normal X allele. FMRP positive villi can be explained by an active normal X allele. The presence of both positive and negative villi indicates that X-inactivation in human CVS is a random process. No villi are found with a mixture of both FMRP-expressing and non-FMRP-expressing cells. This indicates that X-inactivation occurs very early in development, before the villi start to proliferate, and that X-inactivation in villi is a clonal process. In addition, our results indicate that the timing of both X-inactivation and full mutation FMR1 allele inactivation is different, i.e. X-inactivation occurs earlier in development than inactivation of the full mutation.