Long Q et al. (2023), Modification of maternally defined H3K4me3 regu...

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Sci Adv 2023 Apr 07;914:eadd8343. doi: 10.1126/sciadv.add8343.

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Modification of maternally defined H3K4me3 regulates the inviability of interspecific Xenopus hybrids.

Long Q , Yan K , Wang C , Wen Y , Qi F , Wang H , Shi P , Liu X , Chan WY , Lu X , Zhao H .

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Increasing evidence suggests that interspecific hybridization is crucial to speciation. However, chromatin incompatibility during interspecific hybridization often renders this process. Genomic imbalances such as chromosomal DNA loss and rearrangements leading to infertility have been commonly noted in hybrids. The mechanism underlying reproductive isolation of interspecific hybridization remains elusive. Here, we identified that modification of maternally defined H3K4me3 in Xenopus laevis and Xenopus tropicalis hybrids determines the different fates of the two types of hybrids as te×ls with developmental arrest and viable le×ts. Transcriptomics highlighted that the P53 pathway was overactivated, and the Wnt signaling pathway was suppressed in te×ls hybrids. Moreover, the lack of maternal H3K4me3 in te×ls disturbed the balance of gene expression between the L and S subgenomes in this hybrid. Attenuation of p53 can postpone the arrested development of te×ls. Our study suggests an additional model of reproductive isolation based on modifications of maternally defined H3K4me3.

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	Fig. 1. Compensation of the chromosomal deletion in ch3L and ch4L by the homologs of X. tropicalis subgenomes.(A) Embryonic development of the te×ls hybrids (top) and le×ts hybrids (bottom). (B) Germ layer differentiation in wild-type (WT) embryos and hybrid embryos was detected by whole-mount in situ hybridization. Sox17a for endoderm, xbra for mesoderm, and sox2 for ectoderm. le×ts and te×ls are the two types of hybrids. (C and D) Hierarchical clusters of HGEU expression in the comparison of the indicated hybrid versus X. laevis (C) or X. tropicalis (D) at the indicated stages. Individual HGEU values were generated by adding the expression values of the genes located in the lost region in ch3L and ch4L and the corresponding homolog gene from the subgenomes of X. laevis and X. tropicalis. The HGEU value was then normalized to the value of the corresponding HGEU in WT X. laevis (C) or X. tropicalis (D). (E and F) Volcano plots showing fold changes and P values of gene expression in the comparison between le×ts and the embryos of its maternal frog X.l at stage 10 (E) and the gene comparison between te×ls and its maternal frog X.t at stage 10 (F). The x axis represents the fold changes of HGEU, whereas the y axis represents the logarithm of the P value. The P value was generated by unpaired two-tailed t tests. NS, not significant; FC, fold change. X.l, X. laevis; X.t, Xenopus tropicalis.
	Fig. 2. Signaling pathways highlighted in the te×ls hybrids.(A) Dot plots showing the signaling pathways of the DEGs between the te×ls and X. tropicalis embryos from stages 5 to 10. The terms in red highlight denote the cell cycle and DNA damage repair. The terms in blue denote the metabolism pathway. The dot size represents the number of genes enriched by KEGG analysis, and the color of the dots represents the P value generated using the hyper geometric test. CoA, coenzyme A. (B) Ridgeline plot of GSEA between the te×ls hybrids and X. tropicalis embryos at stage 10. A negative value (<0) represents the pathways decreased in the te×ls hybrids, and a positive value (>0) represents the pathways increased in the te×ls hybrids. (C and D) The GSEA analysis of the P53 and Wnt signaling pathways between the te×ls hybrids and X. tropicalis embryos at stage 10. (E and F) Fuzzy c-means clustering identified 10 clusters with distinct temporal patterns of gene expression. The x axis represents the developmental stages in te×ls and X.t, whereas the y axis represents the values of normalized intensity considering the logarithm with base 2 at each stage. (G and H) Heatmap of DEGs involved in the Wnt pathway (G) and DNA damage repair (H) in the comparison of the te×ls hybrids and X. tropicalis from stages 5 to 10 are indicated. X.l, X. laevis; X.t, X. tropicalis.
	Fig. 3. The te×ls hybrids lack maternally defined H3K4me3 modifications.(A to D) Epigenetic modifications of H3K4me3, H3K9ac, and H3K27ac were detected using Western blotting of proteins extracted from the X. laevis (A), le×ts hybrid (B), X. tropicalis (C), and te×ls hybrid (D) embryos at the indicated stages. Histone H3 (H3) was used as the loading control. (E and F) H3K4me3 in the hybrids and WT embryos at stages 5 and 9. (G) Immunofluorescence staining shows H3K4me3 in the hybrids and WT X. tropicalis and X. laevis embryos. Eighteen embryos in each group from three independent experiments were detected. (H) Quantification of the abundance of the H3K4me3 embryos is shown in the left panel (G). At least 204 nuclei of 18 embryos in each group were counted. ***P ≤ 0.001, using one-way analysis of variance (ANOVA) test. X.l, X. laevis; X.t, X. tropicalis.
	Fig. 4. Maternally defined H3K4me3 is involved in regulating differential transcription among the subgenomes in hybrids.(A) Comparison of the ratios of H3k4me3 enrichment in the L chromosome over the S chromosome in the X. laevis, le×ts, and te×ls embryos at stage 9. Wilcoxon rank sum test was used. ***P ≤ 0.001. (B) Overall expression of genes with or without H3K4me3 enrichment in the L, S, and T chromosomes of X. laevis, X. tropicalis, le×ts, and te×ls embryos at stage 9. The gene expression level is represented as the logarithm transcript per million (TPM) to base 2. (C) The correlation between the gene expression ratio and H3K4me3 enrichment ratio of the L to S chromosomes in X. laevis (spearman R = 0.18, P < 2.22 × 10−16). (D) Venn diagram depicting the overlap of the H3K4me3-enriched genes and the genes showing unbalanced expression between the L and S chromosomes in the te×ls hybrids. (E) KEGG pathway analysis of the overlapping genes shown in (D). The dot size represents the number of genes enriched by KEGG analysis, and the color of the dots represents the P value generated using the hyper geometric test. MAPK, mitogen-activated protein kinase. (F) Representative genome views for H3K4me3 ChIP-seq track and transcripts in the X. laevis, le×ts, and te×ls hybrids. mdm2.L, mdm2.S, mdm2.T, rhoa.L, rhoa.S, and rhoa.T. (G) Overexpression of mdm2 extended the development of the te×ls embryos to the neurula stage. (H) Schematic diagram illustrating that maternally defined H3K4me3 modifications regulate reproductive isolation in X. laevis and X. tropicalis, resulting in different fates of the two hybrids. X.l, X. laevis and X.t, X. tropicalis

References [+] :

Akkers, A hierarchy of H3K4me3 and H3K27me3 acquisition in spatial gene regulation in Xenopus embryos. 2009, Pubmed, Xenbase