Pan-family pollen signals control an interspecific stigma barrier across Brassicaceae species.

Prezygotic interspecific incompatibility prevents hybridization between species, which limits interbreeding strategies for crop improvement using wild relatives. The Brassica rapa female self-incompatibility determinant, S-locus receptor kinase (SRK), recognizes interspecific pollen. Here, we report the discovery of a pan-Brassicaceae SRK-interacting interspecific pollen signal (SIPS). On B. rapa stigmas, SIPSs from Arabidopsis and other Brassicaceae species target BrSRK and recruit the female fertility regulator FERONIA receptor kinase to increase stigmatic reactive oxygen species and reduce interspecific pollen viability. Arabidopsis thaliana sips mutant pollen failed to trigger interspecific incompatibility responses. Unlike self-incompatibility, which is controlled by the polymorphic S locus, different genetic variants of SRK interacted comparably with SIPS. This study establishes SIPS-SRK as a Brassicaceae-specific ligand-receptor pair that broadly maintains the stigmatic interspecific barrier in self-incompatible species. Editor's summary: Plant species boundaries are often maintained by reproductive barriers preventing hybridization. Cao et al. identified SIPS, a pollen ligand for the female receptor SRK, and this pair together control compatibility. Whereas a hypervariable region of SRK controls self-incompatibility to prevent self-pollination, SIPS binds to a conserved region of SRK to control interspecies incompatibility. SIPS and SRK homologs are broadly present across the Brassicaceae, indicating that this mechanism maintains multiple species boundaries. —Madeleine Seale INTRODUCTION: Flowering plants have evolved self-incompatibility to specifically prevent mating with close relatives, thereby enforcing outbreeding and avoiding inbred suppression of species vigor. Meanwhile, self-incompatible plants are often challenged by pollen from other species that are nearby and have similar flowering times. Their capacity to enact interspecific incompatibility to reject interspecific pollen, especially from species within the same family, is crucial for maintaining species integrity. However, agriculturally, reproductive barriers established by interspecific incompatibility restrain distant breeding, a key strategy for increasing genetic diversity, exploiting heterosis, and introducing new traits from wild relatives into crops. Despite its far-reaching evolutionary and agricultural importance, the mechanistic basis of interspecific incompatibility remains poorly understood. RATIONALE: In the family Brassicaceae, the female fertility regulator, FERONIA (FER) receptor kinase, maintains a stigmatic barrier of elevated reactive oxygen species (ROS) to reject self-incompatible pollen. The stigma receptor S-locus receptor kinase (SRK) recognizes self-incompatible pollen through an S haplotype–specific interaction with the pollen factor S-locus cysteine rich protein (SCR). This interaction recruits FER and activates ROS to arrest self-incompatible pollen. SRK also recognizes and rejects pollen from other Brassicaceae species on Brassica rapa stigmas. However, the identity of the pollen factor that triggers the interspecific incompatibility responses has remained unknown. RESULTS: We identified SRK-interacting interspecific pollen signal (SIPS) as the pollen factor for interspecific incompatibility. Our results establish that the B. rapa SRK of S46 haplotype (BrSRK46) binds Arabidopsis SIPS1 and SIPS2 with high affinity. Similar to self-incompatibility, this SIPS-SRK complex recruits FER to activate FER-mediated ROS production in the stigma and blocks the hydration and penetration of interspecific pollen into the pistil. On B. rapa stigmas, Arabidopsis thalianasips mutant pollen failed to induce a stigmatic ROS burst, remained viable, successfully hydrated, and overcame the B. rapa stigmatic barrier, demonstrating that SIPS is essential to trigger the interspecific incompatibility response. Evolutionary analyses revealed the broad existence of SIPS across the Brassicaceae family. Although self-incompatible pistils specifically reject self-incompatible pollen, interspecific incompatibility is not S haplotype specific, and pistils with any functional SRK broadly inhibit pollen from the entire Brassicaceae family. This is because, unlike SCRs, SIPSs from distant species are recognized by SRKs irrespective of their S haplotype or species origin. CONCLUSION: Our study demonstrates that the stigmatic receptor SRK interacts with a pan-family interspecific incompatibility pollen factor, forming a SIPS-SRK ligand-receptor pair. This underlies the broad recognition and rejection of interspecific pollen by the stigma of self-incompatible Brassicaceae species. Furthermore, these findings enabled the development of an efficient distant-breeding system to facilitate future efforts to introduce desirable traits from wild relatives into Brassicaceae crops. Model for pan-family SIPS-SRK–triggered interspecific incompatibility in Brassicaceae species.: In interspecific incompatibility, a pan-family, general pollen signal (SIPS) binds various stigma SRKs. In self-incompatibility, an S haplotype–specific pollen signal (SCRx) binds its cognate SRKx (where x represents any S haplotype). Both SIPS-SRK and SCR-SRK interactions promote SRK dimerization, enhance SRK-FER interaction, and activate FER-mediated ROS production. This unified mechanism ultimately inhibits interspecific pollen and self-incompatible pollen on Brassicaceae stigmas. CP, compatible pollen; ISI, interspecific incompatibility; PM, plasma membrane; RBOH, respiratory burst oxidase homolog for ROS production; SI, self-incompatibility. [ABSTRACT FROM AUTHOR]