Tomato clade species (Solanum sect. Lycopersicon) display multiple interspecific reproductive barriers (IRBs). Some IRBs conform to the SI x SC rule, which describes unilateral incompatibility (UI) where pollen from SC species is rejected on SI species’ pistils, but reciprocal pollinations are successful. However, SC x SC UI also exists, offering opportunities to identify factors that contribute to S-RNase-independent IRBs. For instance, SC Solanum pennellii LA0716 pistils only permit SC Solanum lycopersicum pollen tubes to penetrate to the top third of the pistil, while S. pennellii pollen penetrates to S. lycopersicum ovaries. We identified candidate S. pennellii LA0716 pistil barrier genes based on expression profiles and published results. CRISPR/Cas9 mutants were created in eight candidate genes, and mutants were assessed for changes in S. lycopersicum pollen tube growth. Mutants in a gene designated Defective in Induced Resistance 1-like (SpDIR1L), which encodes a small cysteine-rich protein, permitted S. lycopersicum pollen tubes to grow to the bottom third of the style. We show that SpDIR1L protein accumulation correlates with IRB strength and that species with weak or no IRBs toward S. lycopersicum pollen share a 150 bp deletion in the upstream region of SpDIR1L. These results suggest that SpDIR1L contributes to an S-RNase-independent IRB.

Mating system transitions from self-incompatibility (SI) to self-compatibility (SC) are common in plants. In the absence of high levels of inbreeding depression, SC alleles are predicted to spread due to transmission advantage and reproductive assurance. We characterized mating system and pistil-expressed SI factors in 20 populations of the wild tomato species Solanum habrochaites from the southern half of the species range. We found that a single SI to SC transition is fixed in populations south of the Rio Chillon valley in central Peru. In these populations, SC correlated with the presence of the hab-6 S-haplotype that encodes a low activity S-RNase protein. We identified a single population segregating for SI/SC and hab-6. Intrapopulation crosses showed that hab-6 typically acts in the expected codominant fashion to confer SC. However, we found one specific S-haplotype (hab-10) that consistently rejects pollen of the hab-6 haplotype, and results in SI hab-6/hab-10 heterozygotes. We suggest that the hab-10 haplotype could act as a genetic mechanism to stabilize mixed mating in this population by presenting a disadvantage for the hab-6 haplotype. This barrier may represent a mechanism allowing for the persistence of SI when an SC haplotype appears in or invades a population.

A goal of speciation genetics is to understand how the genetic components underlying interspecific reproductive barriers originate within species. Unilateral incompatibility (UI) is a postmating prezygotic barrier in which pollen rejection in the female reproductive tract (style) occurs in only one direction of an interspecific cross. Natural variation in the strength of UI has been observed among populations within species in the wild tomato clade. In some cases, molecular loci underlying self-incompatibility (SI) are associated with this variation in UI, but the mechanistic connection between these intra- and inter-specific pollen rejection behaviors is poorly understood in most instances. We generated an F-2 population between SI and SC genotypes of a single species, Solanum pennellii, to examine the genetic basis of intraspecific variation in UI against other species, and to determine whether loci underlying SI are genetically associated with this variation. We found that F-2 individuals vary in the rate at which UI rejection occurs. One large effect QTL detected for this trait co-localized with the SI-determining S-locus. Moreover, individuals that expressed S-RNase-the S-locus protein involved in SI pollen rejection-in their styles had much more rapid UI responses compared with those without S-RNase protein. Our analysis shows that intraspecific variation at mate choice loci-in this case at loci that prevent self-fertilization-can contribute to variation in the expression of interspecific isolation, including postmating prezygotic barriers. Understanding the nature of such intraspecific variation can provide insight into the accumulation of these barriers between diverging lineages.

In self-incompatible Solanaceae, the pistil protein S-RNase contributes toS-specific pollen rejection in conspecific crosses, as well as to rejecting pollen from foreign species or whole clades. However, S-RNase alone is not sufficient for either type of pollen rejection. We describe a thioredoxin (Trx) type h fromNicotiana alata, NaTrxh, which interacts with and reduces S-RNasein vitro. Here, we show that expressing a redox-inactive mutant, NaTrxh(SS), suppresses bothS-specific pollen rejection and rejection of pollen fromNicotiana plumbaginifolia. Biochemical experiments provide evidence that NaTrxh specifically reduces the Cys(155)-Cys(185)disulphide bond ofS(C10)-Rnase, resulting in a significant increase of its ribonuclease activity. This reduction and increase in S-RNase activity by NaTrxh helps to explain why S-RNase alone could be insufficient for pollen rejection.

Self-incompatibility (SI) mechanisms prevent self-fertilization in flowering plants based on specific discrimination between self- and non-self pollen. Since this trait promotes outcrossing and avoids inbreeding it is a widespread mechanism of controlling sexual plant reproduction. Growers and breeders have effectively exploited SI as a tool for manipulating domesticated crops for thousands of years. However, only within the past thirty years have studies begun to elucidate the underlying molecular features of SI. The specific S-determinants and some modifier factors controlling SI have been identified in the sporophytic system exhibited by Brassica species and in the two very distinct gametophytic systems present in Papaveraceae on one side and in Solanaceae, Rosaceae, and Plantaginaceae on the other. Molecular level studies have enabled SI to SC transitions (and vice versa) to be intentionally manipulated using marker assisted breeding and targeted approaches based on transgene integration, silencing, and more recently CRISPR knock-out of SI-related factors. These scientific advances have, in turn, provided a solid basis to implement new crop production and plant breeding practices. Applications of self-(in)compatibility include widely differing objectives such as crop yield and quality improvement, marker-assisted breeding through SI genotyping, and development of hybrids for overcoming intra- and interspecific reproductive barriers. Here, we review scientific progress as well as patented applications of SI, and also highlight future prospects including further elucidation of SI systems, deepening our understanding of SI-environment relationships, and new perspectives on plant self/non-self recognition.

Abstract Stamens are the male sexual organs in flowers. Their function is to produce and disperse pollen, which carries the plant's sperm cells to the pistil. Mature stamens consist of a filament and an anther. They are initiated on floral meristems and form the third of four concentric organ whorls. The sterile filament provides a vascular connection and positions the anther for pollen dispersal. The anther sits at the tip of the filament and is the site of pollen production. Diploid germ‐line cells in the anther interior undergo meiosis to produce haploid microspores. These microspores, in turn, undergo one or two mitotic divisions, making pollen either bicellular or tricellular at the time of anthesis. Surrounding layers of nonsexual cells are essential for pollen development and also contribute to the external pollen coat. At anthesis, the mature anther splits open, and pollen is available for transfer to the pistil. Key Concepts A canonical flower contains four concentric organ whorls. Stamens comprise the third whorl and function to produce and disperse pollen, which contains the male gametes. Stamens are specified by the combined action of B‐ and C‐class floral homeotic genes in a floral meristem. A mature stamen consists of a nonsexual filament and a four‐lobed anther that is the site of pollen production. Positional information and cell–cell signalling are crucial for specifying the germ‐line pollen mother cells and surrounding somatic tissues. Cells in the interior of the developing anther differentiate into germ‐line archesporial cells and then induce somatic fates in surrounding cells. After meiosis, the germ‐line archesporial cells develop into pollen, and the surrounding somatic cells support pollen development. At anthesis, the mature anther splits and desiccates to allow pollen dispersal.

In plants, transitions in mating system from outcrossing to self-fertilization are common; however, the impact of these transitions on interspecific and interpopulation reproductive barriers is not fully understood. We examined the consequences of mating system transition for reproductive barriers in 19 populations of the wild tomato species Solanum habrochaites. We identified S. habrochaites populations with self-incompatible (SI), self-compatible (SC) and mixed population (MP) mating systems, and characterized pollen-pistil interactions among S. habrochaites populations and between S. habrochaites and other tomato species. We examined the relationship between mating system, floral morphology, interspecific and interpopulation compatibility and pistil SI factors. We documented five distinct phenotypic groups by combining reproductive behavior with molecular data. Transitions from SI to MP were not associated with weakened interspecific reproductive barriers or loss of known pistil SI factors. However, transitions to SC at the northern range margin were accompanied by loss of S-RNase, smaller flowers, and weakened (or absent) interspecific pollen-pistil barriers. Finally, we identified a subset of SC populations that exhibited a partial interpopulation reproductive barrier with central SI populations. Our results support the hypothesis that shifts in mating system, followed by additional loss-of-function mutations, impact reproductive barriers within and between species.

Plants have mechanisms to recognize and reject pollen from other species. Although widespread, these mechanisms are less well understood than the self-incompatibility (SI) mechanisms plants use to reject pollen from close relatives. Previous studies have shown that some interspecific reproductive barriers (IRBs) are related to SI in the Solanaceae. For example, the pistil SI proteins S-RNase and HT protein function in a pistil-side IRB that causes rejection of pollen from self-compatible (SC) red/orange-fruited species in the tomato clade. However, S-RNase-independent IRBs also clearly contribute to rejecting pollen from these species. We investigated S-RNase-independent rejection of Solanum lycopersicum pollen by SC Solanum pennellii LA0716, SC. Solanum habrochaites LA0407, and SC Solanum arcanum LA2157, which lack functional S-RNase expression. We found that all three accessions express HT proteins, which previously had been known to function only in conjunction with S-RNase, and then used RNAi to test whether they also function in S-RNase-independent pollen rejection. Suppressing HT expression in SC S. pennellii LA0716 allows S. lycopersicum pollen tubes to penetrate farther into the pistil in HT suppressed plants, but not to reach the ovary. In contrast, suppressing HT expression in SC. Solanum habrochaites LA0407 and in SC S. arcanum LA2157 allows S. lycopersicum pollen tubes to penetrate to the ovary and produce hybrids that, otherwise, would be difficult to obtain. Thus, HT proteins are implicated in both S-RNase-dependent and S-RNase-independent pollen rejection. The results support the view that overall compatibility results from multiple pollen-pistil interactions with additive effects.

PREMISE OF THE STUDY: Interspecific reproductive barriers (IRBs) often prevent hybridization between closely related species in sympatry. In the tomato clade (Solanum section Lycopersicon), interspecific interactions between natural sympatric populations have not been evaluated previously. In this study, we assessed IRBs between members of the tomato clade from nine sympatric sites in Peru.METHODS: Coflowering was assessed at sympatric sites in Peru. Using previously collected seeds from sympatric sites in Peru, we evaluated premating prezygotic (floral morphology), postmating prezygotic (pollen-tube growth), and postzygotic barriers (fruit and seed development) between sympatric species in common gardens. Pollen-tube growth and seed development were examined in reciprocal crosses between sympatric species.KEY RESULTS: We confirmed coflowering of sympatric species at five sites in Peru. We found three types of postmating prezygotic IRBs during pollen-pistil interactions: (1) unilateral pollen-tube rejection between pistils of self-incompatible species and pollen of self-compatible species; (2) potential conspecific pollen precedence in a cross between two self-incompatible species; and (3) failure of pollen tubes to target ovules. In addition, we found strong postzygotic IRBs that prevented normal seed development in 11 interspecific crosses, resulting in seed-like structures containing globular embryos and aborted endosperm and, in some cases, overgrown endothelium. Viable seed and F-1 hybrid plants were recovered from three of 19 interspecific crosses.CONCLUSIONS: We have identified diverse prezygotic and postzygotic IRBs that would prevent hybridization between sympatric wild tomato species, but interspecific hybridization is possible in a few cases.

PREMISE OF THE STUDY:Self-incompatibility (SI) is a mechanism that prevents inbreeding in many plant species. The mutational breakdown of SI occurs frequently, yet relatively little is known about the evolutionary steps involved in the progressive loss of pistil and pollen SI function.METHODS:In Solanaceae, SI is the S-RNase-based gametophytic type. We used SI and SC populations of the wild tomato species Solanum habrochaites to study natural variation for two pollen SI factors: a Cullin1 (CUL1) protein and an S-locus F-box protein (SLF-23). Pollen compatibility was assessed on an allotriploid tester line encoding an S-RNase recognized by SLF-23. Both pollen factors are required for compatibility on this tester line. Complementation tests and gene sequencing were used to identify mutations in CUL1 or SLF-23.KEY RESULTS:We detected loss-of-function mutations in CUL1 and/or SLF-23 in SC populations collected near the northern and southern geographic margins of this taxon's natural range. Nonmarginal SC and all SI accessions expressed mostly functional alleles of these pollen factors. Comparison of the CUL1 sequences identified several shared deletion mutations present in both northern and southern margin SC accessions.CONCLUSIONS:Loss-of-function mutations in CUL1 and SLF-23 likely became fixed relatively late during SI to SC transitions, after loss of pistil SI function. Mutations in CUL1 establish unilateral incompatibility with SI populations and strengthen reproductive isolation. Point mutations common to northern and southern SC biotypes likely derive from shared ancestral variants found in more central SI populations.

Pollen-pistil interactions contribute to mate selection at the postmating, prezygotic level.

In a new study, the Papaver rhoeas (poppy family) self-incompatibility system has been transferred into Arabidopsis thaliana, a distantly related plant with a very different floral structure. The simple poppy self-incompatibility system may finally make it possible to introduce this potentially valuable trait into any plant.

Two recent studies revealed the genetic architecture of the long fascinating heterostyly in Primula and that the absence of a CYPT gene determines the long style morph.

Involved field radiation therapy (IFRT) alone is potentially curative in stage I-II low-grade follicular lymphoma (FL) but relapse occurs in >50% of cases, predominantly outside RT fields. We hypothesized that systemic therapy given after IFRT would improve disease control, as assessed by progression-free survival (PFS).

PREMISE OF THE STUDY:Interspecific reproductive barriers (IRBs) act to ensure species integrity by preventing hybridization. Previous studies on interspecific crosses in the tomato clade have focused on the success of fruit and seed set. The SI × SC rule (SI species × SC species crosses are incompatible, but the reciprocal crosses are compatible) often applies to interspecific crosses. Because SI systems in the Solanaceae affect pollen tube growth, we focused on this process in a comprehensive study of interspecific crosses in the tomato clade to test whether the SI × SC rule was always followed.METHODS:Pollen tube growth was assessed in reciprocal crosses between all 13 species of the tomato clade using fluorescence microscopy.KEY RESULTS:In crosses between SC and SI species, pollen tube growth follows the SI × SC rule: interspecific pollen tube rejection occurs when SI species are pollinated by SC species, but in the reciprocal crosses (SC × SI), pollen tubes reach ovaries. However, pollen tube rejection occurred in some crosses between pairs of SC species, demonstrating that a fully functional SI system is not necessary for pollen tube rejection in interspecific crosses. Further, gradations in the strength of both pistil and pollen IRBs were revealed in interspecific crosses using SC populations of generally SI species.CONCLUSION:The SI × SC rule explains many of the compatibility relations in the tomato clade, but exceptions occur with more recently evolved SC species and accessions, revealing differences in strength of both pistil and pollen IRBs.

Background: Patients (pts) with metastatic rectal cancer and symptomatic primary, require local and systemic control. Chemotherapy used during chemoradiotherapy (CRT) is adequate for radiosensitisation, but suboptimal for systemic control. The aim of this phase II study was to assess tolerability, local/systemic benefits, of a novel regimen delivering interdigitating intensive chemotherapy with radical CRT. Methods: Eligible pts had untreated synchronous symptomatic primary/metastatic rectal cancer. A total of 12 weeks of treatment with split-course pelvic CRT (total 50.4 Gy with concurrent oxaliplatin and 5-FU infusion) alternating with FOLFOX chemotherapy. All pts staged with CT, MRI and FDG-PET pre and post treatment. Results: Twenty-six pts were treated. Rectal primary MRI stage: T3 81% and T4 15%. Liver metastases in 81%. Twenty-four pts (92%) completed the 12-week regimen. All patients received planned RT dose, and for both agents over 88% of patients achieved a relative dose intensity of >75%. Grade 3 toxicities: neutropenia 23%, diarrhoea 15%, and radiation skin reaction 12%. Grade 4 toxicity: neutropenia 15%. FDG-PET metabolic response rate for rectal primary 96%, and for metastatic disease 60%. Conclusions: Delivery of interdigitating chemotherapy with radical CRT was feasible to treat both primary and metastatic rectal cancer. High completion and response rates were encouraging.