Point Mutations in Centromeric Histone Induce Post-zygotic Incompatibility and Uniparental Inheritance

The centromeric histone 3 variant (CENH3, aka CENP-A) is essential for the segregation of sister chromatids during mitosis and meiosis. To better define CENH3 functional constraints, we complemented a null allele in Arabidopsis with a variety of mutant alleles, each inducing a single amino acid change in conserved residues of the histone fold domain. Many of these transgenic missense lines displayed wild-type growth and fertility on self-pollination, but exhibited frequent post-zygotic death and uniparental inheritance when crossed with wild-type plants. The failure of centromeres marked by these missense mutation in the histone fold domain of CENH3 reproduces the genome elimination syndromes described with chimeric CENH3 and CENH3 from diverged species. Additionally, evidence that a single point mutation is sufficient to generate a haploid inducer provide a simple one-step method for the identification of non-transgenic haploid inducers in existing mutagenized collections of crop species. As proof of the extreme simplicity of this approach to create haploid-inducing lines, we performed an in silico search for previously identified point mutations in CENH3 and identified an Arabidopsis line carrying the A86V substitution within the histone fold domain. This A87V non-transgenic line, while fully fertile on self-pollination, produced postzygotic death and uniparental haploids when crossed to wild type.

着丝粒组蛋白H3变体（centromeric histone 3 variant, CENH3，又名CENP-A）对于有丝分裂与减数分裂过程中姐妹染色单体的分离至关重要。为进一步明确CENH3的功能限制条件，我们采用多种突变等位基因对拟南芥中的无效等位基因进行互补实验，每种突变等位基因均可引发组蛋白折叠结构域（histone fold domain）保守残基发生单氨基酸替换。诸多此类转基因错义突变株系在自交时表现出与野生型一致的生长状态与育性，但在与野生型植株杂交时，会频繁出现合子后死亡与单亲遗传现象。这类携带CENH3组蛋白折叠结构域错义突变的着丝粒所引发的功能缺陷，重现了此前关于嵌合CENH3以及来自趋异物种的CENH3所描述的基因组消除综合征。此外，单点突变即可构建单倍体诱导系（haploid inducer）的相关证据，为从现有作物诱变种质库中筛选非转基因单倍体诱导系提供了一种简便的一步法策略。为验证该方法构建单倍体诱导系的极高简便性，我们通过虚拟筛选（in silico search）查找了此前已报道的CENH3点突变，并筛选到一株拟南芥株系，其组蛋白折叠结构域携带A86V氨基酸替换。该A87V非转基因株系在自交时育性完全正常，但与野生型植株杂交时，会产生合子后死亡与单亲单倍体后代。