Evolution of self-compatibility by a mutant Sm-RNase in citrus

Abstract

Self-incompatibility (SI) is an important mechanism that prevents self-fertilization and inbreeding in flowering plants. The most widespread SI system utilizes S ribonucleases (S-RNases) and S-locus F-boxes (SLFs) as S determinants. In citrus, SI is ancestral, and Citrus maxima (pummelo) is self-incompatible, while Citrus reticulata (mandarin) and its hybrids are self-compatible (SC). Here, we identify nine highly polymorphic pistil-specific, developmentally expressed S-RNases from pummelo that segregate with S haplotypes in a gametophytic manner and cluster with authentic S-RNases. We provide evidence that these S-RNases function as the female S determinants in citrus. Moreover, we show that each S-RNase is linked to approximately nine SLFs. In an analysis of 117 citrus SLF and SFL-like (SLFL) genes, we reveal that they cluster into 12 types and that the S-RNases and intra-haplotypic SLF and SLFL genes co-evolved. Our data support the notion that citrus have a S locus comprising a S-RNase and several SLFs that fit the non-self-recognition model. We identify a predominant single nucleotide mutation, Sm-RNase, in SC citrus, which provides a ‘natural’ loss of function. We show that SI–SC transitions due to the Sm-RNase initially arose in mandarin, spreading to its hybrids and became fixed. Identification of an evolutionarily distant new genus utilizing the S-RNase-based SI system, >100 million years separated from the nearest S-RNase family, is a milestone for evolutionary comparative studies.

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Fig. 1: The citrus S-RNases exhibit key features of S-RNases.
Fig. 2: The pummelo S-RNases exhibit RNase activity and elicit S-specific pollen inhibition in an in vitro bioassay.
Fig. 3: Multiple candidate SLF and SLFL genes are located at the citrus S locus.
Fig. 4: A truncated Sm-RNase appears to be responsible for the loss of SI in citrus.
Fig. 5: Postulated spread of the Sm-RNase and SC in Citrus.
Fig. 6: Phylogenetic relationship of different SI systems in flowering plants.

Data availability

The RNA-seq data shown in Supplementary Table 3 (for pummelo and grapefruit) are available at NCBI BioProject ID under accession codes PRJNA526584 and PRJNA573625. The sequence data shown in Supplementary Table 7 of the pummelo S1-locus and S2-locus BAC clones are available at NCBI BioProject ID under accession codes PRJNA573817 and PRJNA573818, respectively. The DNA sequencing data shown in Supplementary Table 11 from the different citrus species are available at NCBI BioProject ID under accession codes PRJNA544805 (C. maxima), PRJNA544816 (C. aurantium), PRJNA544866 (C. paradisi), PRJNA544867 (C. limon) and PRJNA573624 (C. reticulata). The sequence data of the 15 citrus S-RNase genes are available at NCBI GenBank ID under accession codes MN652897, MN652898, MN652899, MN652900, MN652901, MN652902, MN652903, MN652904, MN652905, MN652906, MN652907, MN652908, MN652909, MN652910, MN652911 and MN652912. Any other raw data are available from the corresponding author upon request.

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Acknowledgements

We are grateful to J. Wu from Nanjing Agricultural University for providing the sample of Pyrus bretschneideri. This research was financially supported by the National Key Research and Development Programme of China (grant no. 2018YFD1000107), the National Natural Science Foundation of China (grant nos. 31772259, 31630065 and 31521092), the Fundamental Research Funds for the Central Universities (grant no. 2662019PY044) and the China Agriculture Research System (grant no. CARS-27). The Biotechnology and Biological Sciences Research Council (BBSRC) funds research in the laboratories of M.B. and V.E.F.-T. (grant no. BB/P005489/1). We would like to thank T. Li (China Agricultural University) and C. Franklin (School of Biosciences, University of Birmingham, UK) for their valuable suggestions.

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L.C., M.L. and V.E.F.-T. designed the experiments. M.L., Z.C., H.Y., Q.X.Jr and M.T. performed the experiments. J.L. completed the collection and sequencing of sour orange. M.L., A.Z., Y. Liu, Y. Li and R.X. analysed the bioinformatics data. S.W., C.C., Z.X. and C.D. collected the pummelo accessions. J.Y., W.G., Q.X., R.M.L., X.D., M.B. and L.C. were involved in the research design and improvement of the manuscript. M.L. and V.E.F.-T. wrote the manuscript.

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Correspondence to Lijun Chai.

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Liang, M., Cao, Z., Zhu, A. et al. Evolution of self-compatibility by a mutant Sm-RNase in citrus. Nat. Plants 6, 131–142 (2020). https://doi.org/10.1038/s41477-020-0597-3

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