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Single-cell allelic HiC analysis, combined with allelic gene expression and chromatin states, reveals parent-of-origin-specific dynamics of chromosome organization and gene expression during mouse preimplantation development.

Paternal and maternal epigenomes undergo marked changes after fertilization(1). Recent epigenomic studies have revealed the unusual chromatin landscapes that are present in oocytes, sperm and early preimplantation embryos, including atypical patterns of histone modifications(2-4) and differences in chromosome organization and accessibility, both in gametes(5-8) and after fertilization(5,8-10). However, these studies have led to very different conclusions: the global absence of local topological-associated domains (TADs) in gametes and their appearance in the embryo(8,9) versus the pre-existence of TADs and loops in the zygote(5,11). The questions of whether parental structures can be inherited in the newly formed embryo and how these structures might relate to allele-specific gene regulation remain open. Here we map genomic interactions for each parental genome (including the X chromosome), using an optimized single-cell high-throughput chromosome conformation capture (HiC) protocol(12,13), during preimplantation in the mouse. We integrate chromosome organization with allelic expression states and chromatin marks, and reveal that higher-order chromatin structure after fertilization coincides with an allele-specific enrichment of methylation of histone H3 at lysine 27. These early parental-specific domains correlate with gene repression and participate in parentally biased gene expression-including in recently described, transiently imprinted loci(14). We also find TADs that arise in a non-parental-specific manner during a second wave of genome assembly. These de novo domains are associated with active chromatin. Finally, we obtain insights into the relationship between TADs and gene expression by investigating structural changes to the paternal X chromosome before and during X chromosome inactivation in preimplantation female embryos(15). We find that TADs are lost as genes become silenced on the paternal X chromosome but linger in regions that escape X chromosome inactivation. These findings demonstrate the complex dynamics of three-dimensional genome organization and gene expression during early development.

Species that propagate by sexual reproduction actively guard against the fertilization of an egg by multiple sperm (polyspermy). Flowering plants rely on pollen tubes to transport their immotile sperm to fertilize the female gametophytes inside ovules. In Arabidopsis, pollen tubes are guided by cysteine-rich chemoattractants to target the female gametophyte(1,2). The FERONIA receptor kinase has a dual role in ensuring sperm delivery and blocking polyspermy(3). It has previously been reported that FERONIA generates a female gametophyte environment that is required for sperm release(4). Here we show that FERONIA controls several functionally linked conditions to prevent the penetration of female gametophytes by multiple pollen tubes in Arabidopsis. We demonstrate that FERONIA is crucial for maintaining de-esterified pectin at the filiform apparatus, a region of the cell wall at the entrance to the female gametophyte. Pollen tube arrival at the ovule triggers the accumulation of nitric oxide at the filiform apparatus in a process that is dependent on FERONIA and mediated by de-esterified pectin. Nitric oxide nitrosates both precursor and mature forms of the chemoattractant LURE1(1), respectively blocking its secretion and interaction with its receptor, to suppress pollen tube attraction. Our results elucidate a mechanism controlled by FERONIA in which the arrival of the first pollen tube alters ovular conditions to disengage pollen tube attraction and prevent the approach and penetration of the female gametophyte by late-arriving pollen tubes, thus averting polyspermy.

FERONIA prevents polyspermy in Arabidopsis by enabling pectin-stimulated nitric oxide accumulation at the filiform apparatus after the first pollen tube arrives, which disengages LURE1 chemoattraction to prevent late-arriving pollen tubes from entering the ovule.