聖誕紅分枝誘導植物菌質體之基因體多樣性及其效應蛋白特性分析

Abstract

聖誕紅分枝誘導性植物菌質體 (以下簡稱聖誕紅菌質體) 普遍存在市面上的聖誕紅品種，可誘發宿主植物產生具園藝價值的多分枝性狀。然而關於聖誕紅分枝性狀的田間穩定性，此類植物菌質體的遺傳多樣性，以及誘發多分枝性狀的機制仍尚未被釐清。為探討以上問題，本研究首先調查十種台灣常見的聖誕紅品種在不同年份的側枝發育數目，以評估聖誕紅多分枝性狀在盆花產業的穩定性。接著，本研究以 Illumina 與 Oxford Nanopore Technologies 等次世代定序技術，針對十五種不同聖誕紅品系所攜帶的菌質體進行基因體解序與比較分析。最後，本研究利用農桿菌於圓葉菸草進行短暫表現試驗，驗證聖誕紅菌質體的效應因子誘導植物側枝增生的功能。根據調查結果，相同聖誕紅品種於四個不同栽培季的側枝數量並不穩定，此現象指出園藝產業對於穩定聖誕紅側枝增生技術的潛在需求。基因體學分析顯示所調查之聖誕紅菌質體菌株皆屬 ‘Candidatus Phytoplasma pruni’，基因體序列平均相似度高達 99.5%。菌株間的低遺傳多樣性可能反應僅由少數個體建立聖誕紅菌質體族群所致的拓荒者效應，或是該菌質體在園藝產業受人為育種選汰的影響。另外，聖誕紅菌質體普遍攜帶兩類具有改變植物性狀潛力的 SAP11 同源效應因子，雖然兩者的胺基酸序列僅有 46.7% 的相似度，但皆能誘導菸草增生相同量級的側枝。總結而言，本研究首次接露聖誕紅菌質體在園藝產業的遺傳多樣性，並證實植物菌質體的效應因子與聖誕紅多分枝性狀的關聯性。未來，藉由釐清參與植物菌質體誘導側枝增生的聖誕紅基因，本研究期望未來能將成果應用於提升聖誕紅育種與盆花生產的效率及穩定性。

Poinsettia branch-inducing (PoiBI) phytoplasmas have been identified as the key factor inducing the free-branching phenotype of poinsettia (Euphorbia pulcherrima) worldwide. Rather than being considered as pathogens, PoiBI phytoplasmas promote compact and bushy growth in poinsettias, thereby enhancing vegetative propagation and commercial value of this ornamental plant. However, knowledge gaps remain regarding the consistency of the poinsettia branching-levels in nurseries, the genomic diversity of PoiBI phytoplasmas, and the molecular mechanism underlying their branch-inducing properties. In this study, we firstly analyzed poinsettia branching data collected across three years in the same growing facility. Next, a metagenomic approach using Illumina and Oxford Nanopore Technologies (ONT) sequencing was employed to study PoiBI phytoplasmas associated with different poinsettia cultivars and their genetic diversity was characterized through comparative genomics. Lastly, an Agrobacterium-mediated transient expression assay in Nicotiana benthamiana was conducted to investigate the functions of putative effector genes. Our results suggested that most poinsettia cultivars exhibited variable branching levels across different years, indicating the need to develop new technologies for stabilizing poinsettia production. In the metagenomic analysis, a total of 15 PoiBI phytoplasma genome assemblies were generated. These genome sequences shared above 99.5 % average nucleotide identity and were all classified as ‘Candidatus Phytoplasma pruni’. The observation of limited genomic variation may be explained by a founder effect at the initial establishment of phytoplasma-poinsettia association or strong selection during cultivar development. Furthermore, among the six conserved putative secreted proteins, two SAP11 homologs sharing only 46.7% amino acid sequence similarity were prevalent among PoiBI phytoplasmas. Despite their sequence divergence, both SAP11 homologs significantly induced branching in N. benthamiana. Overall, this study expanded the genomic catalog of PoiBI phytoplasmas and provided the first evidence linking the free-branching phenotype of poinsettia to specific phytoplasma-encoded effectors. These findings also suggest the potential to improve the efficiency and stability of poinsettia production in the horticultural industry.