Dosage-sensitive maternal siRNAs determine hybridization success in Capsella

Hybrid seed failure arising from wide crosses between plant species is a recurring obstacle in plant breeding, impeding the transfer of desirable traits. This postzygotic reproductive barrier primarily occurs in the endosperm, a tissue that nourishes the embryo and functions similarly to the placenta in mammals. We found that incompatible seeds show a loss of DNA methylation and chromatin condensation in the endosperm, similar to seeds lacking maternal RNA polymerase IV activity. This similarity is linked to a decrease in small interfering RNAs in the endosperm (sirenRNAs), maternal RNA polymerase IV-dependent short interfering RNAs that regulate DNA methylation. Several AGAMOUS-like MADS-box transcription factor genes (AGLs), key regulators of endosperm development, are targeted by sirenRNAs in cis and in trans. This finding aligns with the enrichment of AGL target genes among deregulated genes. We propose that hybrid seed failure results from reduced maternal sirenRNAs combined with increased AGL expression, leading to abnormal gene regulation in the endosperm. Each RNA-Seq library was prepared from seeds taken out of 10-15 siliques. Three biological replicates were used for each genotype. The BS-Seq and sRNA-SEq libraries were prepared from manually dissected endosperm of around 600 seeds. Two biological replicates were used for each genotype.种间远缘杂交引发的杂交种子败育是植物育种中反复出现的障碍，阻碍了优良性状的转移。这种合子后生殖障碍主要发生在胚乳——一种为胚胎提供营养、功能类似哺乳动物胎盘的组织——中。我们发现，不亲和杂交种子的胚乳中存在DNA甲基化缺失与染色质浓缩异常现象，与缺失母本RNA聚合酶IV（RNA polymerase IV）活性的种子表现一致。该相似性与胚乳中小干扰RNA（small interfering RNAs，sirenRNAs）的水平降低相关——这类由母本RNA聚合酶IV依赖生成的短干扰RNA可调控DNA甲基化。多个AGAMOUS样MADS-box转录因子基因（AGAMOUS-like MADS-box transcription factor genes，缩写为AGLs）作为胚乳发育的关键调控因子，可被sirenRNAs通过顺式和反式作用进行靶向调控。该发现与失调基因中AGL靶基因的富集现象相符。我们提出，杂交种子败育源于母本sirenRNAs水平降低与AGLs表达上调共同作用，最终引发胚乳内基因调控异常。每份RNA测序（RNA-Seq）文库均从10~15个角果中取出的种子制备得到，每个基因型设置3次生物学重复。亚硫酸氢盐测序（BS-Seq）与小RNA测序（sRNA-Seq）文库则从约600粒经手工解剖的胚乳中制备，每个基因型设置2次生物学重复。