Flowering plants have evolved numerous intraspecific and interspecific prezygotic reproductive barriers to prevent production of unfavourable offspring¹. Within a species, self-incompatibility (SI) is a widely utilized mechanism that rejects self-pollen^2,3 to avoid inbreeding depression. Interspecific barriers restrain breeding between species and often follow the SI × self-compatible (SC) rule, that is, interspecific pollen is unilaterally incompatible (UI) on SI pistils but unilaterally compatible (UC) on SC pistils^1,4–6. The molecular mechanisms underlying SI, UI, SC and UC and their interconnections in the Brassicaceae remain unclear. Here we demonstrate that the SI pollen determinant S-locus cysteine-rich protein/S-locus protein 11 (SCR/SP11)^2,3 or a signal from UI pollen binds to the SI female determinant S-locus receptor kinase (SRK)^2,3, recruits FERONIA (FER)^7–9 and activates FER-mediated reactive oxygen species production in SI stigmas^10,11 to reject incompatible pollen. For compatible responses, diverged pollen coat protein B-class^12–14 from SC and UC pollen differentially trigger nitric oxide, nitrosate FER to suppress reactive oxygen species in SC stigmas to facilitate pollen growth in an intraspecies-preferential manner, maintaining species integrity. Our results show that SRK and FER integrate mechanisms underlying intraspecific and interspecific barriers and offer paths to achieve distant breeding in Brassicaceae crops.

Understanding adaptive genetic variation and whether it can keep pace with predicted future climate change is critical in assessing the genetic vulnerability of species and developing conservation management strategies. The lack of information on adaptive genetic variation in relict species carrying abundant genetic resources hinders the assessment of genetic vulnerability. Using a landscape genomics approach, this study aimed to determine how adaptive genetic variation shapes population divergence and to predict the adaptive potential of Pterocarya macroptera (a vulnerable relict species in China) under future climate scenarios. We applied restriction site-associated DNA sequencing (RAD-seq) to obtain 8244 single-nucleotide polymorphisms (SNPs) from 160 individuals across 28 populations. We examined the pattern of genetic diversity and divergence, and then identified outliers by genetic differentiation (F_ST) and genotype–environment association (GEA) methods. We further dissected the effect of geographical/environmental gradients on genetic variation. Finally, we predicted genetic vulnerability and adaptive risk under future climate scenarios. We identified three genetic lineages within P. macroptera: the Qinling-Daba-Tianmu Mountains (QDT), Western Sichuan (WS) and Northwest Yunnan (NWY) lineages, which showed significant signals of isolation by distance (IBD) and isolation by environment (IBE). IBD and IBE explained 3.7–5.7 and 8.6–12.8 % of the genetic structure, respectively. The identified GEA SNP-related genes were involved in chemical defence and gene regulation and may exhibit higher genetic variation to adapt to the environment. Gradient forest analysis revealed that the genetic variation was mainly shaped by temperature-related variables, indicating its adaptation to local thermal environments. A limited adaptive potential was suggested by the high levels of genetic vulnerability in marginal populations. Environmental gradient mainly shaped the population differentiation of P. macroptera. Marginal populations may be at high risk of extinction, and thus proactive management measures, such as assisted gene flow, are required to ensure the survival of these populations.

Differences in local abundance and ploidy level are predicted to impact the direction of introgression between species. Here, we tested these hypotheses on populations of Betula albosinensis (red birch) and Betula platyphylla (white birch) which were thought to differ in ploidy level, the former being tetraploid and the latter diploid. We sampled 391 birch individuals from nine localities in China, and classified them into species based on leaf morphology. Twelve nuclear microsatellite markers were genotyped in each sample, and analysed using principal coordinates analysis and STRUCTURE software. We compared the effects of two different methods of scoring polyploid genotypes on population genetic analyses. We analysed the effect of ploidy, local species abundance and latitude on levels of introgression between the species. Leaf morphology divided our samples into red and white birch, but genetic analyses unexpectedly revealed two groups within red birch, one of which was tetraploid, as expected, but the other of which appeared to have diploid microsatellite genotypes. Five individuals were identified as early-generation hybrids or backcrosses between white birch and red birch and five were identified between red birch and ‘diploid’ red birch. Cline analysis showed that levels of admixture were not significantly correlated with latitude. Estimated genetic differentiation among species was not significantly different between determined tetraploid and undetermined tetraploid genotypes. Limited hybridization and gene flow have occurred between red birch and white birch. Relative species abundance and ploidy level do not impact the direction of introgression between them, as genetic admixture is roughly symmetrical. We unexpectedly found populations of apparently diploid red birch and this taxon may be a progenitor of allotetraploid red birch populations. Incomplete lineage sorting may explain patterns of genetic admixture between apparently diploid and allotetraploid red birch.

Delineating closely related and morphologically similar species is difficult. Here, we integrate morphology, genetics, ploidy and geography to resolve species and subspecies boundaries in four trees of section Costatae (genus Betula): Betula ashburneri, B. costata, B. ermanii and B. utilis, as well as multiple subspecies and polyploid races. We genotyped 371 individuals (20–133 per species) from 51 populations at 15 microsatellite markers, as well as a subset of individuals, using restriction-site associated DNA sequencing and nuclear internal transcribed spacers. We determined the ploidy level of eight individuals using flow cytometry and characterized leaf variation for a subset of 109 individuals by morphometric analysis. Integration of multiple lines of evidence suggested a series of revisions to the taxonomy of section Costatae. Betula costata and B. ermanii were found to be valid. Molecular and leaf morphology analyses revealed little differentiation between diploid B. albosinensis and some samples of B. utilis ssp. utilis. By contrast, other B. utilis ssp. utilis samples and ssp. albosinensis formed a morphological continuum but differed based on genetics. Specifically, B. utilis ssp. albosinensis was divided into two groups with group I genetically similar to B. utilis ssp. utilis and group II, a distinct cluster, proposed as the new diploid species Betula buggsii sp. nov. Phylogenomic analysis based on 2285 620 single nucleotide polymorphisms identified a well-supported monophyletic clade of B. buggsii. Morphologically, B. buggsii is characterized by elongated lenticels and a distinct pattern of bark peeling and may be geographically restricted to the Qinling–Daba Mountains. Our integrated approach identifies six taxa within section Costatae: B. ashburneri, B. buggsii, B. costata, B. utilis ssp. utilis, B. utilis ssp. albosinensis and B. ermanii. Our research demonstrates the value of an integrative approach using morphological, geographical, genetic and ploidy-level data for species delineation.