Haldane's rule in the placenta: sex-biased misregulation of the Kcnq1 imprinting cluster in hybrid mice

Hybrid phenotypes that contribute to postzygotic reproductive isolation often exhibit pronounced asymmetry, both between reciprocal crosses and between the sexes in accordance with Haldane's rule. Inviability in mammalian hybrids is associated with parent-of-origin placental growth abnormalities for which misregulation of imprinted genes is the leading candidate mechanism. However, direct evidence for the involvement of imprinted genes in hybrid growth dysplasia is limited. We used transcriptome and reduced representation bisulfite sequencing to conduct the first genome-scale assessment of the contribution of imprinted genes to parent-of-origin placental growth dysplasia in the cross between the house mouse (Mus musculus domesticus) and the Algerian mouse (Mus spretus). Imprinted genes with transgressive expression and methylation were concentrated in the Kcnq1 cluster, which contains causal genes for prenatal growth abnormalities in mice and humans. Hypermethylation of the cluster's imprinting control region, and consequent misexpression of the genes Phlda2 and Ascl2, is a strong candidate mechanism for transgressive placental undergrowth. Transgressive placental and gene regulatory phenotypes, including expression and methylation in the Kcnq1 cluster, were more extreme in hybrid males. While consistent with Haldane's rule, male-biased defects are unexpected in rodent placenta because the X-chromosome is effectively hemizygous in both sexes. In search of an explanation we found evidence of leaky imprinted (paternal) X-chromosome inactivation in hybrid female placenta, an epigenetic disturbance that may buffer females from the effects of X-linked incompatibilities to which males are fully exposed. Sex differences in chromatin structure on the X and sex-biased maternal effects are non-mutually exclusive alternative explanations for adherence to Haldane's rule in hybrid placenta. The results of this study contribute to understanding the genetic basis of hybrid inviability in mammals, and the role of imprinted genes in speciation. Overall design: Analysis of 29 samples of placental DNA at e17.5 of the same pregnancies (3 groups, 4-5 biological replicates per sex, per group)

参与后合子生殖隔离（postzygotic reproductive isolation）的杂交表型通常呈现显著的不对称性，该不对称性既存在于正反交之间，也符合霍尔丹法则（Haldane's rule）所描述的性别间差异模式。哺乳动物杂交个体的致死性与亲本起源型胎盘生长异常密切相关，其中印记基因（imprinted genes）的调控紊乱是目前最具潜力的候选机制。然而，关于印记基因参与杂交个体生长发育异常的直接证据仍较为有限。本研究利用转录组测序与简化代表性亚硫酸氢盐测序（reduced representation bisulfite sequencing），首次在全基因组层面评估了印记基因在家鼠（Mus musculus domesticus）与阿尔及利亚鼠（Mus spretus）的杂交后代中，对亲本起源型胎盘生长异常的贡献。表达与甲基化水平出现越轨的印记基因富集于Kcnq1基因簇（Kcnq1 cluster），该基因簇包含小鼠与人类产前生长异常的致病基因。该基因簇的印记控制区域（imprinting control region）发生高甲基化，进而导致Phlda2与Ascl2基因表达紊乱，这是引发胎盘越轨性发育不全的有力候选机制。杂交雄性个体的胎盘越轨表型与基因调控异常（包括Kcnq1基因簇的表达与甲基化异常）更为显著。尽管这一结果符合霍尔丹法则，但啮齿类胎盘中出现雄性偏向的发育缺陷却出乎意料——因为两性的X染色体均处于功能性半合子状态。为探寻成因，我们在杂交雌性胎盘中发现了印记松弛型（父本型）X染色体失活（X-chromosome inactivation）的证据；这种表观遗传紊乱可能使雌性免受X连锁不相容性的影响，而雄性则完全暴露于该效应之下。X染色体染色质结构的性别差异与性别偏向性母源效应，均可作为杂交胎盘中符合霍尔丹法则的非互斥备选解释。本研究结果有助于解析哺乳动物杂交个体致死性的遗传基础，以及印记基因在物种形成过程中的作用。实验整体设计：对妊娠第17.5天（e17.5）的29份胎盘DNA样本进行分析，共分为3组，每组每性别设置4-5个生物学重复。