Genomic analysis of sex chromosomes and sex differentiation in Caricaceae

Papaya is an excellent model species to study mechanisms for sex determination and sex chromosome evolution in plants, due to the presence of three sex types: female, male, and hermaphrodite. Previous research has revealed the location of the sex determination region (SDR) and the existence of three sex chromosomes X, Y, and Yh in the papaya genome, including the number and identity of transcripts within this region. It is still unknown which gene or genes in the SDR are responsible for sex determination in papaya. The regulatory mechanism for sex determination and expression in papaya remains a mystery despite that multiple candidate genes have been proposed. This Dissertation focuses on three different molecular approaches to search for candidate genes responsible for sex determination in C. papaya and to elucidate possible regulatory mechanisms for sex expression in papaya flowers. The first approach included a transcriptome analysis of papaya flower buds from different sex types at different developmental stages, including the transcriptome analysis of hermaphrodite-to-female sex reversal mutants. The analysis of differentially expressed genes between male, female, and hermaphrodite flower buds (Chapter 2) revealed the existence of several genes with strong sex-biased expression in male and hermaphrodite flowers that might be playing an important role in male organ development, specifically tapetum development in the anther developmental pathway. Other transcription factors related to flower organ development and flowering time regulation, as well as genes that are known to participate in ABA, ROS and auxin signaling pathways were identified as differentially expressed among papaya flowers. The analysis of differentially expressed genes between hermaphrodite-to-female and hermaphrodite flower buds (Chapter 3) revealed only five differentially expressed genes. Among these genes, only an uncharacterized gene with a high level of expression in hermaphrodite-to-female mutants and female flower buds was identified. Despite no match for this gene was found in other plant species, the function of this gene might be related to biological processes involved in the anther developmental pathway. None of these differentially expressed genes located in the SDR in papaya, it is possible that these genes act downstream sex determination genes as consequence of a complex regulatory cascade, regulated by a Y-specific gene acting as a stamen promoting factor. A second approach involved identifying differences on the promoter region of genes located in the sex chromosomes (X, Y and Yh) in papaya and studying the effects of these differences on the expression of a previously proposed candidate gene for sex determination called CpMDAR4 (Chapter 4). Results showed that differences in the promoter sequence of the CpMDAR4 gene drove the expression of a reporter gene (GUS) to different flower tissues in Arabidopsis thaliana, due to the existence and location of cis-regulatory elements (POLLEN1LELAT52) responsible for anther or pollen-specific expression present on the Y chromosomes (Y and Yh). A similar expression pattern of CpMDAR4 alleles in different tissues of papaya flowers and the existence of two different isoforms for this gene on male and hermaphrodites support MDAR4 as an important candidate for sex differentiation. Sex chromosomes are hypothesized to have evolved from the same pair of autosomes in the Caricaceae family, therefore a third approach involved the genome assembly of a species from a closely related genus (Jarilla heterophylla) and identification of conserved synteny in the SDR between both species (Chapter 5). Conserved candidate genes in the SDR between the two species (C. papaya and J. heterophylla) are more likely to be responsible for sex determination given the close phylogenetic relationship, overlapping geographical distribution in southern Mexico, and similar morphological and reproductive characteristics between these species. Results showed that J. heterophylla has a XY sex determination system, with homomorphic sex chromosomes, as was found in papaya, and genes in the SDR are conserved between these species. Only one Y-specific gene in papaya (CpY19 or CpSVP) was identified in J. heterophylla, making this gene an important candidate for sex differentiation in papaya.LimitedAuthor requested closed access (OA after 2yrs) in Vireo ETD syste