Self-incompatibility Phenotype Research Articles

Transposable elements cause the loss of self-incompatibility in citrus.

Self-incompatibility (SI) is a widespread prezygotic mechanism for flowering plants to avoid inbreeding depression and promote genetic diversity. Citrus has an S-RNase-based SI system, which was frequently lost during evolution. We previously identified a single nucleotide mutation in Sm-RNase, which is responsible for the loss of SI in mandarin and its hybrids. However, little is known about other mechanisms responsible for conversion of SI to self-compatibility (SC) and we identify a completely different mechanism widely utilized by citrus. Here, we found a 786-bp miniature inverted-repeat transposable element (MITE) insertion in the promoter region of the FhiS2-RNase in Fortunella hindsii Swingle (a model plant for citrus gene function), which does not contain the Sm-RNase allele but are still SC. We demonstrate that this MITE plays a pivotal role in the loss of SI in citrus, providing evidence that this MITE insertion prevents expression of the S-RNase; moreover, transgenic experiments show that deletion of this 786-bp MITE insertion recovers the expression of FhiS2-RNase and restores SI. This study identifies the first evidence for a role for MITEs at the S-locus affecting the SI phenotype. A family-wide survey of the S-locus revealed that MITE insertions occur frequently adjacent to S-RNase alleles in different citrus genera, but only certain MITEs appear to be responsible for the loss of SI. Our study provides evidence that insertion of MITEs into a promoter region can alter a breeding strategy and suggests that this phenomenon may be broadly responsible for SC in species with the S-RNase system.

Identification of a novel self-compatible phenotype and its mutant SRK allele harbourin an LTR-retrotransposon in radish (Raphanus sativus L.)

ABSTRACT Self-incompatibility (SI) responses of radish (Raphanus sativus L.) are determined by two tightly linked genes encoding an S receptor kinase (SRK) and an S-locus cysteine-rich protein/S locus protein 11 (SCR/SP11), respectively. A radish showing an almost self-compatible (SC) phenotype was identified in this study. Inheritance patterns showed that this SC phenotype was dominant over an SI phenotype. In addition, the SC phenotype co-segregated with an S haplotype in an F2 population. This SC radish contained an RsS-26 haplotype in which duplicate SRK-like genes were previously identified. Full-length sequences of two SRK-like genes of 18,133-bp and 6,200-bp in length were obtained, and they were designated as RsSRK-26-1 and RsSRK-26-2, respectively. No critical mutation was found in the coding region of SRK-like genes. However, a 4,146-bp intact LTR-retrotransposon was identified in the third intron of RsSRK-26-1 of the SC radish. Interestingly, this LTR-retrotransposon was not detected in three other breeding lines containing the same RsS-26 haplotype. While normal transcription of RsSRK-26-1 and SI phenotypes were observed in these three breeding lines, no transcript of RsSRK-26-1 was detected in the SC radish, suggesting that recent transposition of an LTR-retrotransposon in the RsSRK-26-1 gene might be responsible for the SC phenotype of radish.

Challenges and Perspectives in the Study of Self-Incompatibility in Orchids.

Self-incompatibility affects not only the formation of seeds, but also the evolution of species diversity. A robust understanding of the molecular mechanisms of self-incompatibility is essential for breeding efforts, as well as conservation biology research. In recent years, phenotypic and multiple omics studies have revealed that self-incompatibility in Orchidaceae is mainly concentrated in the subfamily Epidendroideae, and the self-incompatibility phenotypes are diverse, even in the same genus, and hormones (auxin and ethylene), and new male and female determinants might be involved in SI response. This work provides a good foundation for future studies of the evolution and molecular mechanisms of self-incompatibility. We review recent research progress on self-incompatibility in orchids at the morphological, physiological, and molecular levels, provide a general overview of self-incompatibility in orchids, and propose future research directions.

Genetic mapping of sex and self-incompatibility determinants in the androdioecious plant Phillyrea angustifolia

The diversity of mating and sexual systems in angiosperms is spectacular, but the factors driving their evolution remain poorly understood. In plants of the Oleaceae family, an unusual self-incompatibility (SI) system has been discovered recently, whereby only two distinct homomorphic SI specificities segregate stably. To understand the role of this peculiar SI system in preventing or promoting the diversity of sexual phenotypes observed across the family, an essential first step is to characterize the genetic architecture of these two traits. Here, we developed a high-density genetic map of the androdioecious shrub P. angustifolia based on a F1 cross between a hermaphrodite and a male parent with distinct SI genotypes. Using a double restriction-site associated digestion (ddRAD) sequencing approach, we obtained reliable genotypes for 196 offspring and their two parents at 10,388 markers. The resulting map comprises 23 linkage groups totaling 1,855.13 cM on the sex-averaged map. We found strong signals of association for the sex and SI phenotypes, that were each associated with a unique set of markers on linkage group 12 and 18 respectively, demonstrating inheritance of these traits as single, independent, mendelian factors. The P. angustifolia linkage map shows robust synteny to the olive tree genome overall. Two of the six markers strictly associated with SI in P. angustifolia have strong similarity with a recently identified 741kb chromosomal region fully linked to the SI phenotype on chromosome 18 of the olive tree genome, providing strong cross-validation support. The SI locus stands out as being markedly rearranged, while the sex locus has remained relatively more collinear between the two species. This P. angustifolia linkage map will be a useful resource to investigate the various ways by which the sex and SI determination systems have co-evolved in the broader phylogenetic context of the Oleaceae family.

Characterization of a Common S Haplotype BnS-6 in the Self-Incompatibility of Brassica napus

Self-incompatibility (SI) is a pollen-stigma recognition system controlled by a single and highly polymorphic genetic locus known as the S-locus. The S-locus exists in all Brassica napus (B. napus, AACC), but natural B. napus accessions are self-compatible. About 100 and 50 S haplotypes exist in Brassica rapa (AA) and Brassica oleracea (CC), respectively. However, S haplotypes have not been detected in B. napus populations. In this study, we detected the S haplotype distribution in B. napus and ascertained the function of a common S haplotype BnS-6 through genetic transformation. BnS-1/BnS-6 and BnS-7/BnS-6 were the main S haplotypes in 523 B. napus cultivars and inbred lines. The expression of SRK in different S haplotypes was normal (the expression of SCR in the A subgenome affected the SI phenotype) while the expression of BnSCR-6 in the C subgenome had no correlation with the SI phenotype in B. napus. The BnSCR-6 protein in BnSCR-6 overexpressed lines was functional, but the self-compatibility of overexpressed lines did not change. The low expression of BnSCR-6 could be a reason for the inactivation of BnS-6 in the SI response of B. napus. This study lays a foundation for research on the self-compatibility mechanism and the SI-related breeding in B. napus.

Characterization of self-incompatible Brassica napus lines lacking SP11 expression.

Recognition of self-incompatibility (SI) is regulated by the SRK and SP11 genes in Brassicaceae. Brassica rapa and B. oleracea are self-incompatible, while most cultivated species of B. napus, which arose from hybridization between B. rapa and B. oleracea, are self-compatible. Various studies of the SRK and SP11 genes in self-compatible B. napus have been reported, but details of the mechanism in different B. napus lines are not fully understood. In this study, we confirmed the S haplotypes, SI phenotypes and SP11 expression in 10 representative lines of B. napus, and identified two SI lines (N110 and N343) lacking SP11 expression. In N343 (with BnS1 and BnS6 haplotypes), we confirmed that there is a 3.6-kb insertion in the promoter region of BnSP11-1, and that BnSP11-1 and BnSP11-6 are not expressed, as reported previously (expression of BnSP11-6 is suppressed by the BnS1 haplotype), although this line is self-incompatible. Similarly, in N110, with two novel S haplotypes (BnS8 and BnS9) in addition to BnS6, a 4.3-kb insertion was identified in the promoter region of BnSP11-9, and expression levels of BnSP11-6, BnSP11-8 and BnSP11-9 were all suppressed (BnSP11-6 and BnSP11-8 may be suppressed by BnS8 and BnS9, respectively), although the phenotype was self-incompatible. This observation of an SI phenotype without SP11 expression suggests the existence of unknown factor(s) that induce pollen-stigma incompatibility in B. napus.

Evolution of self-incompatibility in the Brassicaceae: Lessons from a textbook example of natural selection.

Self‐incompatibility (SI) is a self‐recognition genetic system enforcing outcrossing in hermaphroditic flowering plants and results in one of the arguably best understood forms of natural (balancing) selection maintaining genetic variation over long evolutionary times. A rich theoretical and empirical population genetics literature has considerably clarified how the distribution of SI phenotypes translates into fitness differences among individuals by a combination of inbreeding avoidance and rare‐allele advantage. At the same time, the molecular mechanisms by which self‐pollen is specifically recognized and rejected have been described in exquisite details in several model organisms, such that the genotype‐to‐phenotype map is also pretty well understood, notably in the Brassicaceae. Here, we review recent advances in these two fronts and illustrate how the joint availability of detailed characterization of genotype‐to‐phenotype and phenotype‐to‐fitness maps on a single genetic system (plant self‐incompatibility) provides the opportunity to understand the evolutionary process in a unique perspective, bringing novel insight on general questions about the emergence, maintenance, and diversification of a complex genetic system.

Impact of whole genome triplication on the evolutionary history and the functional dynamics of regulatory genes involved in Brassica self-incompatibility signalling pathway.

Polyploidy or whole genome duplication is a frequent and recurrent phenomenon in flowering plants that has played a major role in their diversification, adaptation and speciation. The adaptive success of polyploids relates to the different evolutionary fates of duplicated genes. In this study, we explored the impact of the whole genome triplication (WGT) event in the Brassiceae tribe on the genes involved in the self-incompatibility (SI) signalling pathway, a mechanism allowing recognition and rejection of self-pollen in hermaphrodite plants. By taking advantage of the knowledge acquired on this pathway as well as of several reference genomes in Brassicaceae species, we determined copy number of the different genes involved in this pathway and investigated their structural and functional evolutionary dynamics. We could infer that whereas most genes involved in the SI signalling returned to single copies after the WGT event (i.e. ARC1, JDP1, THL1, THL2, Exo70A01) in diploid Brassica species, a few were retained in duplicated (GLO1 and PLDα) or triplicated copies (MLPK). We also carefully studied the gene structure of these latter duplicated genes (including the conservation of functional domains and active sites) and tested their transcription in the stigma to identify which copies seem to be involved in the SI signalling pathway. By taking advantage of these analyses, we then explored the putative origin of a contrasted SI phenotype between two Brassica rapa varieties that have been fully sequenced and shared the same S-allele (S60).

Phylogenetic analysis of S-RNase-like genes from Fragaria provides evidence for ancestral S-RNase duplication in the Rosaceae lineage

Self-incompatibility (SI) is associated with cultural and breeding difficulties in rosaceous fruit trees such as cherry, plum, apple, and pear. Many species in Rosaceae, Solanaceae, and Plantaginaceae have S-RNase-based gametophytic SI, with S-RNase and F-box proteins as pistil S and pollen S determinants, respectively. Although the S-RNase-based gametophytic SI system is generally believed to have evolved once in a eudicot ancestor before the divergence of asterids and rosids, we previously found that S-RNase and its homologs were duplicated in the ancestral genome of Rosaceae. Here, we reinvestigated the evolutionary paths of S-RNase-like genes using homologs from the diploid Fragaria species F. nipponica (SI), F. nubicola (SI), and F. iinumae (SI phenotype undetermined). Our phylogenetic analysis and estimation of proxy ages for the establishment of S-RNase and its homologs in rosaceous species supports the previously inferred ancestral S-RNase duplication in the Rosaceae lineage. The divergence of S-RNases in subtribe Malinae and Prunus predated the most recent common ancestor of Rosaceae, which includes genes from three rosaceous species (Fragaria, Prunus, and Malus). The results of the present study strengthen the idea that the genus Prunus and subtribe Malinae have evolved different S-RNases for SI recognition. These results may explain the genus- or subtribe-specific SI recognition mechanism operating in rosaceous fruit trees and will be valuable for artificial SI control and self-compatible breeding.

Plasticity of the self-incompatibility phenotype inBrassica oleracea

Plasticity of the self-incompatibility phenotype inBrassica oleracea

Morphological Type Identification of Self-Incompatibility in Dendrobium and Its Phylogenetic Evolution Pattern.

Self-incompatibility (SI) is a type of reproductive barrier within plant species and is one of the mechanisms for the formation and maintenance of the high diversity and adaptation of angiosperm species. Approximately 40% of flowering plants are SI species, while only 10% of orchid species are self-incompatible. Intriguingly, as one of the largest genera in Orchidaceae, 72% of Dendrobium species are self-incompatible, accounting for nearly half of the reported SI species in orchids, suggesting that SI contributes to the high diversity of orchid species. However, few studies investigating SI in Dendrobium have been published. This study aimed to address the following questions: (1) How many SI phenotypes are in Dendrobium, and what are they? (2) What is their distribution pattern in the Dendrobium phylogenetic tree? We investigated the flowering time, the capsule set rate, and the pollen tube growth from the representative species of Dendrobium after artificial pollination and analysed their distribution in the Asian Dendrobium clade phylogenetic tree. The number of SI phenotypes exceeded our expectations. The SI type of Dendrobium chrysanthum was the primary type in the Dendrobium SI species. We speculate that there are many different SI determinants in Dendrobium that have evolved recently and might be specific to Dendrobium or Orchidaceae. Overall, this work provides new insights and a comprehensive understanding of Dendrobium SI.

Examination of S-Locus Regulated Differential Expression in Primula vulgaris Floral Development.

Recent findings on the molecular basis of heteromorphic self-incompatibility in Primula have shown that the controlling self-incompatibility (S)-locus is not allelic, but is instead a small hemizygous region of only a few genes in the thrum genotype. How these genes alter the development of floral morphology and the specificity of self-incompatibility is still not completely clear. In order to start to identify genes regulated by the S-locus and elucidate the large-scale biological processes affected, we used RNA-seq data from floral buds of heteromorphic P. vulgaris pin (long style, short anthers) and thrum (short style, long anthers) morphs at early and late developmental time points. Differential expression between the two morphs was assessed at both time points and Gene Ontology term analyses of these gene sets were conducted. Our findings suggest that the S-locus regulates a large number of genes outside its physical bounds and likely sets up a cascade of expression changes. Additionally, we found evidence to suggest that there may be a timing difference in pollen development between the morphs, with pin pollen development proceeding earlier than thrum pollen development. This finding provides insight into how morphological differences in pollen between the morphs may be established, but intriguingly, could also be related to the self-incompatibility phenotype.

Commonalities and differences between Brassica and Arabidopsis self-incompatibility.

In higher plants, the self-incompatibility mechanism is important for inhibition of self-fertilization and facilitation of out-crossing. In Brassicaceae, the self-incompatibility response is mediated by allele-specific interaction of the stigma-localized S-locus receptor kinase (SRK) with the pollen coat-localized ligand (SCR/SP11). All self-incompatible Brassicaceae plants analyzed have been found to have the SRK and SCR/SP11 genes in the S-locus region. Although Arabidopsis thaliana is self-compatible, transformation with functional SRK-SCR genes from self-incompatible Arabidopsis species confers the self-incompatibility phenotype to A. thaliana. The allele-specific interaction between SRK and SCR activates the downstream signaling cascade of self-incompatibility. Yeast two-hybrid analysis with a kinase domain of SRK as bait and genetic analysis suggested several candidate components of self-incompatibility signaling in Brassica. Recently, A. thaliana genes orthologous to the identified genes for Brassica self-incompatibility signaling were evaluated by using a self-incompatible transgenic A. thaliana plant and these orthologous genes were found not to be involved in self-incompatibility signaling in the transgenic A. thaliana. In this review, we describe common and different aspects of S-locus genomic regions and self-incompatibility signaling between Brassica and Arabidopsis.

The ARC1 E3 Ligase Promotes a Strong and Stable Self-Incompatibility Response in Arabidopsis Species: Response to the Nasrallah and Nasrallah Commentary

Following the identification of the male (S-locus Cysteine Rich/S-locus Protein 11) and female (S Receptor kinase [SRK]) factors controlling self-incompatibility in the Brassicaceae, research in this field has focused on understanding the nature of the cellular responses activated by these regulators. We previously identified the ARM Repeat Containing1 (ARC1) E3 ligase as a component of the SRK signaling pathway and demonstrated ARC1's requirement in the stigma for self-incompatible pollen rejection in Brassica napus, Arabidopsis lyrata, and Arabidopsis thaliana. Here, we discuss our findings on the role of ARC1 in reconstructing a strong and stable A. thaliana self-incompatibility phenotype, in the context of the putative issues outlined in a commentary by Nasrallah and Nasrallah. Additionally, with their proposed standardized strategy for studying self-incompatibility in A. thaliana, we offer our perspective on what constitutes a strong and stable self-incompatibility phenotype in A. thaliana and how this should be investigated and reported to the greater community.

Robust self-incompatibility in the absence of a functional ARC1 gene in Arabidopsis thaliana.

Self-incompatibility (SI) is the primary determinant of the outbreeding mode of sexual reproduction in the Brassicaceae. All Arabidopsis thaliana accessions analyzed to date carry mutations that disrupt SI functions by inactivating the SI specificity-determining S locus or SI modifier loci. S-locus genes isolated from self-incompatible close relatives of A. thaliana restore robust SI in several accessions that harbor only S-locus mutations and confer transient SI in accessions that additionally harbor mutations at modifier loci. Self-incompatible transgenic A. thaliana plants have proved to be valuable for analysis of the recognition and signaling events that underlie SI in the Brassicaceae. Here, we review results demonstrating that S-locus genes are necessary and sufficient for SI signaling and for restoration of a strong and developmentally stable SI phenotype in several accessions of A. thaliana. The data indicate that introduction of a functional E3 ligase-encoding ARC1 gene, which is deleted in all accessions that have been analyzed to date, is not required for SI signaling leading to inhibition of self pollen or for reversion of A. thaliana to its fully self-incompatible ancestral state.

High-density genetic maps for loci involved in nuclear male sterility (NMS1) and sporophytic self-incompatibility (S-locus) in chicory (Cichorium intybus L., Asteraceae).

High-density genetic maps were constructed for loci involved in nuclear male sterility (NMS1-locus) and sporophytic self-incompatibility (S-locus) in chicory (Cichorium intybus L.). The mapping population consisted of 389 F1' individuals derived from a cross between two plants, K28 (male-sterile) and K59 (pollen-fertile), both heterozygous at the S-locus. This F1' mapping population segregated for both male sterility (MS) and strong self-incompatibility (SI) phenotypes. Phenotyping F1' individuals for MS allowed us to map the NMS1-locus to linkage group (LG) 5, while controlled diallel and factorial crosses to identify compatible/incompatible phenotypes mapped the S-locus to LG2. To increase the density of markers around these loci, bulked segregant analysis was used. Bulks and parental plants K28 and K59 were screened using amplified fragment length polymorphism (AFLP) analysis, with a complete set of 256 primer combinations of EcoRI-ANN and MseI-CNN. A total of 31,000 fragments were generated, of which 2,350 showed polymorphism between K59 and K28. Thirteen AFLP markers were identified close to the NMS1-locus and six in the vicinity of the S-locus. From these AFLP markers, eight were transformed into sequence-characterized amplified region (SCAR) markers and of these five showed co-dominant polymorphism. The chromosomal regions containing the NMS1-locus and the S-locus were each confined to a region of 0.8cM. In addition, we mapped genes encoding proteins similar to S-receptor kinase, the female determinant of sporophytic SI in the Brasicaceae, and also markers in the vicinity of the putative S-locus of sunflower, but none of these genes or markers mapped close to the chicory S-locus.

Regulation of the S-Locus Receptor Kinase and Self-Incompatibility inArabidopsis thaliana

Intraspecific mate selectivity often is enforced by self-incompatibility (SI), a barrier to self-pollination that inhibits productive pollen-pistil interactions. In the Brassicaceae, SI specificity is determined by two highly-polymorphic proteins: the stigmatic S-locus receptor kinase (SRK) and its pollen coat-localized ligand, the S-locus cysteine-rich protein (SCR). Arabidopsis thaliana is self fertile, but several of its accessions can be made to express SI, albeit to various degrees, by transformation with functional SRK-SCR gene pairs isolated from its close self-incompatible relative, Arabidopsis lyrata. Here, we use a newly identified induced mutation that suppresses the SI phenotype in stigmas of SRK-SCR transformants of the Col-0 accession to investigate the regulation of SI and the SRK transgene. This mutation disrupts NRPD1a, a gene that encodes a plant-specific nuclear RNA polymerase required for genomic methylation and production of some types of silencing RNAs. We show that NRPD1a, along with the RNA-dependent RNA polymerase RDR2, is required for SI in some A. thaliana accessions. We also show that Col-0 nrpd1a mutants exhibit decreased accumulation of SRK transcripts in stigmas, which is not, however, responsible for loss of SI in these plants. Together, our analysis of the nrpd1a mutation and of SRK promoter activity in various accessions reveals that the SRK transgene is subject to several levels of regulation, which vary substantially by tissue type and by accession. This study thus helps explain the well-documented differences in expression of SI exhibited by SRK-SCR transformants of different A. thaliana accessions.

SELF-INCOMPATIBILITY IN OLIVE: A NEW HYPOTHESIS ON THE S-LOCUS GENES CONTROLLING POLLEN-PISTIL INTERACTION

SELF-INCOMPATIBILITY IN OLIVE: A NEW HYPOTHESIS ON THE S-LOCUS GENES CONTROLLING POLLEN-PISTIL INTERACTION

Self-sterility in flowering plants: preventing self-fertilization increases family diversification rates

New data are presented on the distribution and frequency of self-sterility (SS) - predominantly pre-zygotic self-incompatibility (SI) systems - in flowering plants and the hypothesis is tested that families with self-sterile taxa have higher net diversification rates (DRs) than those with exclusively self-compatible taxa using both absolute and relative rate tests. Three major forms of SI systems (where pollen is rejected at the stigmatic, stylar or ovarian interface) are found to occur in the oldest families of flowering plants, with times of divergence >100 million years before the present (mybp), while post-fertilization SS and heterostyly appear in families with crown ages of 81 and 87 mybp, respectively. It is also founnd that many (22) angiosperm families exhibit >1 SI phenotype and that the distribution of different types of SS does not show strong phylogenetic clustering, collectively suggesting that SS and SI systems have evolved repeatedly de novo in angiosperm history. Families bearing self-sterile taxa have higher absolute DRs using all available calibrations of the angiosperm tree, and this affect is caused mostly by the high DR of families with homomorphic SI systems (in particular stigmatic SI) or those in which multiple SS/SI phenotypes have been observed (polymorphic). Lastly, using sister comparisons, it is further demonstrated that in 29 of 38 sister pairs (including 95 families), the self-sterile sister group had higher species richness and DR than its self-compatible sister based on either the total number of taxa in the clade with SS or only the estimated fraction to harbour SS based on literature surveys. Collectively, these analyses point to the importance of SS, particularly pre-zygotic SI in the evolution of flowering plants.

Evolutionary genetics of an S-like polymorphism in Papaveraceae with putative function in self-incompatibility.

Background Papaver rhoeas possesses a gametophytic self-incompatibility (SI) system not homologous to any other SI mechanism characterized at the molecular level. Four previously published full length stigmatic S-alleles from the genus Papaver exhibited remarkable sequence divergence, but these studies failed to amplify additional S-alleles despite crossing evidence for more than 60 S-alleles in Papaver rhoeas alone.Methodology/Principal FindingsUsing RT-PCR we identified 87 unique putative stigmatic S-allele sequences from the Papaveraceae Argemone munita, Papaver mcconnellii, P. nudicuale, Platystemon californicus and Romneya coulteri. Hand pollinations among two full-sib families of both A. munita and P. californicus indicate a strong correlation between the putative S-genotype and observed incompatibility phenotype. However, we also found more than two S-like sequences in some individuals of A. munita and P. californicus, with two products co-segregating in both full-sib families of P. californicus. Pairwise sequence divergence estimates within and among taxa show Papaver stigmatic S-alleles to be the most variable with lower divergence among putative S-alleles from other Papaveraceae. Genealogical analysis indicates little shared ancestral polymorphism among S-like sequences from different genera. Lack of shared ancestral polymorphism could be due to long divergence times among genera studied, reduced levels of balancing selection if some or all S-like sequences do not function in incompatibility, population bottlenecks, or different levels of recombination among taxa. Preliminary estimates of positive selection find many sites under selective constraint with a few undergoing positive selection, suggesting that self-recognition may depend on amino acid substitutions at only a few sites.Conclusions/SignificanceBecause of the strong correlation between genotype and SI phenotype, sequences reported here represent either functional stylar S-alleles, tightly linked paralogs of the S-locus or a combination of both. The considerable complexity revealed in this study shows we have much to learn about the evolutionary dynamics of self-incompatibility systems.