Sequence and chromatin characteristics of the largest known plant sex chromosome

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Unisexuality of land plant sporophytes has evolved many times, implying the independent origin of various forms of sex determination, including XY sex determination, with either homomorphic sex chromosomes or heteromorphic sex chromosomes. The latter are known from Cannabis, Humulus, Silene, Rumex, and Coccinia, the Cucurbitaceae that is the focus of the present project, continuing the PI's work on unveiling stages of sex chromosome evolution in plants and the evolution of dimorphic sex chromosomes in the genus Coccinia. We have confirmed that the Y chromosome of C. grandis is the largest in land plants, while the species' total genome is relatively small and phylogenetically close to Cucumis, a genus with >5 fully annotated genomes of important crops. The present work will address two gaps in our understanding of plant sex chromosomes: (i) lack of analysis of the DNA composition of heteromorphic sex chromosomes, (ii) lack of information on whether X and Y chromosomes show histone differences, as is the case in dosage-compensated animal XY systems. To fill these gaps, we will isolate sex chromosomes by flow cytometry and microdissection, and generate Illumina-sequenced low-coverage genomes of a male plant, a female plant, pure X chromosomes, and pure Y chromosomes. Bioinformatics analyses will then reveal the types and relative frequencies of the repetitive DNA that is expected to underlie the huge size increase in the Y chromosome compared to the autosomes and X chromosome. Relevant work pipelines for Illumina data were established in the PI's lab during (on repetitive DNA and polyploidy) and will be adapted for the present project. Pilot data obtained by the postdoc to be working in this project during visits to the labs of J. Fuchs and A. Houben, IPK, Gatersleben, show that the X and the Y chromosome can be microdissected and that Illumina sequencing of isolated chromosomes is feasible. Likewise, pilot data on histone methylation suggest sex-specific patterns. Lastly, using repetitive DNA-based new probes, FISH in Coccinia grandis and related species that we have in cultivation will clarify whether other Coccinia species have incipient heteromorphic sex chromosomes or not.