CHK1-CENP B/MAD2 is associated with mild oxidative damage-induced sex chromosome aneuploidy of male mouse embryos during in vitro fertilization

Abstract A high incidence of aneuploidy is observed in vitro fertilization (IVF)-derived embryos, but the formation and repair mechanisms are unknown. Here, we investigated the effects of slightly increased reactive oxygen species (ROS) produced by in vitro culture conditions on embryo aneuploidy and the roles of the spindle assembly checkpoint (SAC) protein, mitotic arrest-deficient 2 (MAD2), and the DNA damage response (DDR) protein, checkpoint kinase 1 (CHK1), in aneuploidy repair. By assessing chromosome abnormalities via DAPI staining, karyotype analysis and next-generation sequencing technology, we demonstrated that mild oxidative damage mainly increased the risk of sex chromosome aneuploidy in male mouse embryos (41,XXY,+X and 41,XYY,+Y) through chromosome mis-segregation during the first mitosis. Isobaric tags for relative and absolute quantitation technology revealed that mild oxidative damage inhibited the expression of male reproduction-related proteins, including a kinase anchor protein 4 (AKAP4), whose gene is located on mouse/human Chromosome X. Under mild oxidative damage, abrogation of MAD2 by MAD2 inhibitor-1 (M2I-1) or CHK1 by siRNA microinjection increased sex chromosome mosaicism rate and reduced mitosis-promoting factor (MPF) activity. CHK1 inhibition also reduced kinetochore localization of centromere protein B (CENP B) and MAD2. These findings show that DDR and SAC are responsible for repair of sex chromosome mosaicism via the pCHK1 (S345)-CENP B/MAD2-MPF pathway; further, IVF may have negative effects on male offspring's reproduction ability, which ultimately depends on their continued repair capability. Therefore, we suggest that antioxidants, especially those targeting improved CHK1-MAD2 function, may be a promising therapeutic strategy to reduce aneuploidy formation of IVF-derived embryos and to maintain genome integrity of embryo and offspring.