FAFU and UIUC Joint Center for Genomics and Biotechnology, Key Laboratory of Sugarcane Biology and Genetic Breeding Ministry of Agriculture, Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Fujian Agriculture and Forestry University, Fuzhou 350002, Fujian, China.
Department of Plant Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Plant Sci. 2018 Jul;272:99-106. doi: 10.1016/j.plantsci.2018.04.001. Epub 2018 Apr 5.
Papaya is trioecious and an excellent system for studying sex determination and differentiation in plants. An ortholog of HUA1, CpHUA1, a gene controlling stamen and carpel development in Arabidopsis, was cloned and characterized in papaya. CpHUA1 consists of 12 exons with full genomic length of 19,313 bp in male AU9 and 19,312 bp in hermaphrodite SunUp, whereas the Arabidopsis HUA1 consists of 12 exons with full genomic length of 4300 bp. All the 324 SNPs between male and hermaphrodite varieties are in the 11th intron, which spans 8.5 kb. Quantitative RT-PCR revealed that CpHUA1 expression is highly elevated in carpels, suggesting that CpHUA1 may be involved in sex differentiation gene network. Southern blot analysis revealed a distinct restriction pattern in male AU9 compared to hermaphrodite Kapoho and SunUp, despite high DNA sequence identity and sharing of all but two EcoR I restriction sites in genomic CpHUA1 sequences of AU9 and SunUp. The methylation of cytosine at one restriction site in male but not in other two sex types may result in distinct restriction pattern of EcoR I in southern blot result. Bisulfite sequencing showed differential methylation of CpHUA1 among sex types, particularly the enrichment of sex-specific methylation in 9th and 11th intron. The methylation difference in cold stress induced male to hermaphrodite mutant mostly observed in the CHH context of CpHUA1, but no methylation difference detected in CHH context in other sex types, which may indicate the role of methylation in CHH context of CpHUA1 in temperature-related stress response and sex reversal.
番木瓜是三性花，是研究植物性别决定和分化的极好系统。拟南芥中控制雄蕊和心皮发育的 HUA1 的同源基因 CpHUA1 在番木瓜中被克隆并进行了特征描述。CpHUA1 由 12 个外显子组成，在雄性 AU9 中的全长基因组为 19313bp，在两性 SunUp 中的全长基因组为 19312bp，而拟南芥 HUA1 由 12 个外显子组成，全长基因组为 4300bp。雄性和两性品种之间的所有 324 个 SNP 都在第 11 个内含子中，该内含子跨越 8.5kb。定量 RT-PCR 显示 CpHUA1 在雌蕊中的表达高度上调，表明 CpHUA1 可能参与性别分化基因网络。Southern blot 分析显示，尽管 AU9 和 SunUp 的基因组 CpHUA1 序列在 DNA 序列同一性和除两个 EcoR I 限制位点之外的所有限制位点共享，但 AU9 与两性 Kapoho 和 SunUp 相比，雄性 AU9 中存在明显的限制模式。一个限制位点处胞嘧啶的甲基化可能导致 EcoR I 在 Southern blot 结果中的限制模式不同。亚硫酸氢盐测序显示，CpHUA1 在不同性别类型之间存在差异甲基化，特别是在第 9 个和第 11 个内含子中存在性别特异性甲基化的富集。在冷胁迫诱导的雄性到两性突变体中观察到 CpHUA1 中 CHH 上下文的差异甲基化，但在其他性别类型中未检测到 CHH 上下文的甲基化差异，这可能表明 CHH 上下文的甲基化在温度相关应激反应和性别逆转中发挥作用。
