Supplementary Material for: Nucleolar Dominance and Repression of 45S Ribosomal RNA Genes in Hybrids between Xenopus borealis and X. muelleri (2n = 36)

Nucleolar dominance is a dramatic disruption in the formation of nucleoli and the expression of ribosomal RNA (rRNA) genes, characteristic of some plant and animal hybrids. Here, we report that F1 hybrids produced from reciprocal crosses between 2 sister species of Xenopus clawed frogs, X. muelleri and X. borealis, undergo nucleolar dominance somewhat distinct from a pattern previously reported in hybrids between phylogenetically more distant Xenopus species. Patterns of nucleolar development, 45S rRNA expression, and gene copy inheritance were investigated using a combination of immunostaining, pyrosequencing, droplet digital PCR, flow cytometry, and epigenetic inhibition. In X. muelleri × X. borealis hybrids, typically only 1 nucleolus is formed, and 45S rRNA genes are predominantly expressed from 1 progenitor's alleles, X. muelleri, regardless of the cross-direction. These changes are accompanied by an extensive (∼80%) loss of rRNA gene copies in the hybrids relative to their parents, with the transcriptionally underdominant variant (X. borealis) being preferentially lost. Chemical treatment of hybrid larvae with a histone deacetylase inhibitor resulted in a partial derepression of the underdominant variant. Together, these observations shed light on the genetic and epigenetic basis of nucleolar dominance as an underappreciated manifestation of genetic conflicts within a hybrid genome.核仁显性（nucleolar dominance）是指核仁形成与核糖体RNA（rRNA）基因表达发生显著紊乱的现象，常见于部分动植物杂交种。本研究针对爪蟾属（Xenopus）的两个姐妹种——穆勒爪蟾（X. muelleri）与北方爪蟾（X. borealis）的正反交F1杂交种展开分析，发现其核仁显性模式与此前在亲缘关系更远的爪蟾物种杂交种中报道的模式存在一定差异。研究人员通过免疫染色（immunostaining）、焦磷酸测序（pyrosequencing）、微滴式数字PCR（droplet digital PCR）、流式细胞术（flow cytometry）以及表观遗传抑制实验等多种手段，对核仁发育模式、45S rRNA表达情况以及基因拷贝数遗传模式进行了探究。在穆勒爪蟾×北方爪蟾杂交种中，通常仅形成1个核仁，且45S rRNA基因主要由穆勒爪蟾的等位基因表达，这一规律不受正反交方向影响。相较于亲本，杂交种的rRNA基因拷贝数出现了显著丢失（约80%），其中转录活性呈显性不足的北方爪蟾等位基因拷贝发生了优先丢失。对杂交幼体施加组蛋白去乙酰化酶抑制剂（histone deacetylase inhibitor）处理后，显性不足的北方爪蟾等位基因的表达得到了部分去阻遏。综上，本研究的观测结果揭示了核仁显性的遗传与表观遗传基础——这是一种长期以来未被充分重视的杂交基因组（hybrid genome）内遗传冲突（genetic conflicts）的表现形式。