Nicotiana suaveolens accessions with different ploidy levels exhibit different reproductive isolation mechanisms in interspecific crosses with Nicotiana tabacum.

Reproductive isolation, including prezygotic and postzygotic barriers, is a mechanism that separates species. Many species in the Nicotiana section Suaveolentes exhibit reproductive isolation in crosses with Nicotiana tabacum. In this study, we investigated whether the chromosome numbers and ploidy levels of eight Nicotiana suaveolens accessions are related to the reproductive isolation after crosses with N. tabacum by flow cytometry and chromosome analyses. Additionally, the internal transcribed spacer (ITS) regions of the eight N. suaveolens accessions were sequenced and compared with the previously reported sequences of 22 Suaveolentes species to elucidate the phylogenetic relationships in the section Suaveolentes. We revealed that four N. suaveolens accessions comprised 64 chromosomes, while the other four accessions carried 32 chromosomes. Depending on the ploidy levels of N. suaveolens, several types of reproductive isolation were observed after crosses with N. tabacum, including decreases in the number of capsules and the germination rates of hybrid seeds, as well as hybrid lethality and abscission of enlarged ovaries at 12-17 days after pollination. A phylogenetic analysis involving ITS sequences divided the eight N. suaveolens accessions into three distinct clades. Based on the results, we confirmed that N. suaveolens accessions vary regarding ploidy levels and reproductive isolation mechanisms in crosses with N. tabacum. These accessions will be very useful for revealing and characterizing the reproductive isolation mechanisms in interspecific crosses and their relationships with ploidy levels.

Parental Genome Imbalance Causes Hybrid Seed Lethality as Well as Ovary Abscission in Interspecific and Interploidy Crosses in Nicotiana.

Enhanced ovary abscission after pollination and hybrid seed lethality result in post-zygotic reproductive isolation in plant interspecific crosses. However, the connection between these barriers remains unclear. Here, we report that an imbalance in parental genomes or endosperm balance number (EBN) causes hybrid seed lethality and ovary abscission in both interspecific and intraspecific-interploidy crosses in the genus Nicotiana. Auxin treatment suppressed ovary abscission, but not hybrid seed lethality, in an interspecific cross between Nicotiana suaveolens and N. tabacum, suggesting that ovary abscission-related genes are located downstream of those involved in hybrid seed lethality. We performed interploidy crosses among N. suaveolens tetraploids, octoploids, and neopolyploids and revealed hybrid seed lethality and ovary abscission in interploid crosses. Furthermore, a higher maternal EBN than paternal EBN caused these barriers, as previously observed in N. suaveolens × N. tabacum crosses. Altogether, these results suggest that maternal excess of EBN causes hybrid seed lethality, which in turn leads to ovary abscission through the same mechanism in both interspecific and interploidy crosses.