S-RNase Alleles Associated With Self-Compatibility in the Tomato Clade: Structure, Origins, and Expression Plasticity.

The self-incompatibility (SI) system in the Solanaceae is comprised of cytotoxic pistil S-RNases which are countered by S-locus F-box (SLF) resistance factors found in pollen. Under this barrier-resistance architecture, mating system transitions from SI to self-compatibility (SC) typically result from loss-of-function mutations in genes encoding pistil SI factors such as S-RNase. However, the nature of these mutations is often not well characterized. Here we use a combination of S-RNase sequence analysis, transcript profiling, protein expression and reproductive phenotyping to better understand different mechanisms that result in loss of S-RNase function. Our analysis focuses on 12 S-RNase alleles identified in SC species and populations across the tomato clade. In six cases, the reason for gene dysfunction due to mutations is evident. The six other alleles potentially encode functional S-RNase proteins but are typically transcriptionally silenced. We identified three S-RNase alleles which are transcriptionally silenced under some conditions but actively expressed in others. In one case, expression of the S-RNase is associated with SI. In another case, S-RNase expression does not lead to SI, but instead confers a reproductive barrier against pollen tubes from other tomato species. In the third case, expression of S-RNase does not affect self, interspecific or inter-population reproductive barriers. Our results indicate that S-RNase expression is more dynamic than previously thought, and that changes in expression can impact different reproductive barriers within or between natural populations.

Transcriptome Profiling and Genome-Wide Association Studies Reveal GSTs and Other Defense Genes Involved in Multiple Signaling Pathways Induced by Herbicide Safener in Grain Sorghum.

Herbicide safeners protect cereal crops from herbicide injury by inducing genes and proteins involved in detoxification reactions, such as glutathione S-transferases (GSTs) and cytochrome P450s (P450s). Only a few studies have characterized gene or protein expression profiles for investigating plant responses to safener treatment in cereal crops, and most transcriptome analyses in response to safener treatments have been conducted in dicot model species that are not protected by safener from herbicide injury. In this study, three different approaches were utilized in grain sorghum (Sorghum bicolor (L.) Moench) to investigate mechanisms involved in safener-regulated signaling pathways. An initial transcriptome analysis was performed to examine global gene expression in etiolated shoot tissues of hybrid grain sorghum following treatment with the sorghum safener, fluxofenim. Most upregulated transcripts encoded detoxification enzymes, including P450s, GSTs, and UDP-dependent glucosyltransferases (UGTs). Interestingly, several of these upregulated transcripts are similar to genes involved with the biosynthesis and recycling/catabolism of dhurrin, an important chemical defense compound, in these seedling tissues. Secondly, 761 diverse sorghum inbred lines were evaluated in a genome-wide association study (GWAS) to determine key molecular-genetic factors governing safener-mediated signaling mechanisms and/or herbicide detoxification. GWAS revealed a significant single nucleotide polymorphism (SNP) associated with safener-induced response on chromosome 9, located within a phi-class SbGST gene and about 15-kb from a different phi-class SbGST. Lastly, the expression of these two candidate SbGSTs was quantified in etiolated shoot tissues of sorghum inbred BTx623 in response to fluxofenim treatment. SbGSTF1 and SbGSTF2 transcripts increased within 12-hr after fluxofenim treatment but the level of safener-induced expression differed between the two genes. In addition to identifying specific GSTs potentially involved in the safener-mediated detoxification pathway, this research elucidates a new direction for studying both constitutive and inducible mechanisms for chemical defense in cereal crop seedlings.

Transcriptomic analysis links gene expression to unilateral pollen-pistil reproductive barriers.

BACKGROUND: Unilateral incompatibility (UI) is an asymmetric reproductive barrier that unidirectionally prevents gene flow between species and/or populations. UI is characterized by a compatible interaction between partners in one direction, but in the reciprocal cross fertilization fails, generally due to pollen tube rejection by the pistil. Although UI has long been observed in crosses between different species, the underlying molecular mechanisms are only beginning to be characterized. The wild tomato relative Solanum habrochaites provides a unique study system to investigate the molecular basis of this reproductive barrier, as populations within the species exhibit both interspecific and interpopulation UI. Here we utilized a transcriptomic approach to identify genes in both pollen and pistil tissues that may be key players in UI. RESULTS: We confirmed UI at the pollen-pistil level between a self-incompatible population and a self-compatible population of S. habrochaites. A comparison of gene expression between pollinated styles exhibiting the incompatibility response and unpollinated controls revealed only a small number of differentially expressed transcripts. Many more differences in transcript profiles were identified between UI-competent versus UI-compromised reproductive tissues. A number of intriguing candidate genes were highly differentially expressed, including a putative pollen arabinogalactan protein, a stylar Kunitz family protease inhibitor, and a stylar peptide hormone Rapid ALkalinization Factor. Our data also provide transcriptomic evidence that fundamental processes including reactive oxygen species (ROS) signaling are likely key in UI pollen-pistil interactions between both populations and species. CONCLUSIONS: Gene expression analysis of reproductive tissues allowed us to better understand the molecular basis of interpopulation incompatibility at the level of pollen-pistil interactions. Our transcriptomic analysis highlighted specific genes, including those in ROS signaling pathways that warrant further study in investigations of UI. To our knowledge, this is the first report to identify candidate genes involved in unilateral barriers between populations within a species.

Interspecific reproductive barriers between sympatric populations of wild tomato species (Solanum section Lycopersicon).

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 F1 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.

Testing the SI × SC rule: Pollen-pistil interactions in interspecific crosses between members of the tomato clade (Solanum section Lycopersicon, Solanaceae).

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.

Interspecific reproductive barriers in the tomato clade: opportunities to decipher mechanisms of reproductive isolation.

The tomato clade within the genus Solanum has numerous advantages for mechanistic studies of reproductive isolation. Its thirteen closely related species, along with four closely allied Solanum species, provide a defined group with diverse mating systems that display complex interspecific reproductive barriers. Several kinds of pre- and postzygotic barriers have already been identified within this clade. Well-developed genetic maps, introgression lines, interspecific bridging lines, and the newly available draft genome sequence of the domesticated tomato (Solanum lycopersicum) are valuable tools for the genetic analysis of interspecific reproductive barriers. The excellent chromosome morphology of these diploid species allows detailed cytological analysis of interspecific hybrids. Transgenic methodologies, well developed in the Solanaceae, allow the functional testing of candidate reproductive barrier genes as well as live imaging of pollen rejection events through the use of fluorescently tagged proteins. Proteomic and transcriptomics approaches are also providing new insights into the molecular nature of interspecific barriers. Recent progress toward understanding reproductive isolation mechanisms using these molecular and genetic tools is assessed in this review.