Varkonyi-Gasic Erika, Wang Tianchi, Cooney Janine, Jeon Subin, Voogd Charlotte, Douglas Mikaela J, Pilkington Sarah M, Akagi Takashi, Allan Andrew C
The New Zealand Institute for Plant and Food Research Ltd (PFR), Private Bag 92169, Auckland, 1142, New Zealand.
The New Zealand Institute for Plant and Food Research Ltd (PFR), Hamilton, 3240, New Zealand.
New Phytol. 2021 May;230(4):1461-1475. doi: 10.1111/nph.17234. Epub 2021 Feb 28.
Kiwifruit (Actinidia chinensis) is a dioecious, long-living woody perennial vine. Reduced generation time and induction of hermaphroditism can accelerate crop improvement and facilitate alternative farming for better food security in the face of climate change. Previous studies identified that CENTRORADIALIS genes CEN and CEN4 act to repress flowering, whilst the male-specific Shy Girl (SyGl) gene with homology to type-C cytokinin response regulators could repress gynoecium development in model plants. Here we use CRISPR/Cas9 to mutagenize CEN, CEN4 and SyGl in the male kiwifruit A. chinensis 'Bruce'. Biallelic mutations of CEN and CEN4 generated rapid-flowering male plants, and simultaneous targeting of CEN4 and SyGl gave rise to rapid-flowering hermaphrodites with restored gynoecial function and viable pollen, providing functional evidence for the role of SyGl in suppression of feminization. Analysis of ovary tissues identified genes that contribute to carpel development and revealed that SyGl affected both cytokinin profiles and the expression of genes involved in cytokinin metabolism and signalling. The plant lines generated by CEN4/SyGl knockout could self-pollinate and produce fast-flowering offspring. These results establish that SyGI acts as the suppressor of feminization in kiwifruit and demonstrate the potential for accelerated breeding in an outcrossing horticultural woody perennial.
猕猴桃（中华猕猴桃）是一种雌雄异株的多年生木质藤本植物，寿命较长。缩短世代时间并诱导雌雄同株可以加速作物改良，并在面对气候变化时促进替代种植方式，以实现更好的粮食安全。先前的研究表明，辐射状中心基因CEN和CEN4具有抑制开花的作用，而与C类细胞分裂素响应调节因子具有同源性的雄性特异性害羞女孩（SyGl）基因可以抑制模式植物中的雌蕊发育。在此，我们使用CRISPR/Cas9对雄性猕猴桃中华猕猴桃‘布鲁诺’中的CEN、CEN4和SyGl进行诱变。CEN和CEN4的双等位基因突变产生了开花迅速的雄性植株，同时靶向CEN4和SyGl则产生了开花迅速的雌雄同株植株，其雌蕊功能得以恢复且花粉可育，这为SyGl在抑制雌性化中的作用提供了功能证据。对卵巢组织的分析确定了有助于心皮发育的基因，并揭示SyGl既影响细胞分裂素谱，也影响参与细胞分裂素代谢和信号传导的基因表达。通过敲除CEN4/SyGl产生的植株系能够自花授粉并产生开花迅速的后代。这些结果表明，SyGI是猕猴桃雌性化的抑制因子，并证明了在异交园艺多年生木本植物中加速育种的潜力。
