利用代謝體學探討由植物細胞激素AtCAPE9調控的免疫反應

Abstract

系統性後天抗性 (Systemic acquired resistance, SAR) 是植物在局部病原體感染後啟動的全株防禦反應，主要由水楊酸 (Salicylic acid, SA) 和致病蛋白一號 (Pathogenesis-related protein 1, PR1) 調控。然而，SAR中系統性信號的傳遞機制仍不明確。在之前的研究中，我們發現PR1會被酵素切割產生植物內生性胜肽AtCAPE9，AtCAPE9在由細菌鞭毛蛋白衍生胜肽flg22誘導的局部和系統免疫反應中發揮了關鍵調節作用。儘管AtCAPE9已被證明是SA信號途徑中的重要分子，但其在調控SAR中的下游機制仍不清楚。為了瞭解這一機制，我們透過液相層析串聯式質譜儀 (LC-MS/MS) 對經AtCAPE9處理的阿拉伯芥葉片和韌皮部滲出液進行了代謝體學分析。結果顯示，初級和次級代謝物均發生顯著變化，其中N-羥基哌啶酸 (N-hydroxypipecolic acid, NHP) 和核黃素 (Riboflavin) 在葉片和韌皮部滲出液中均顯著增加，表明其可能在局部和系統免疫中具有重要作用。進一步的功能研究證實，NHP在AtCAPE9誘導的免疫中是不可或缺的，因為缺乏將哌啶酸 (Pip) 轉化為NHP的黃素依賴單加氧酶1 (Flavin-dependent monooxygenase 1, FMO1) 的阿拉伯芥突變體無法表現AtCAPE9誘導的局部和系統免疫。此外，作為NHP信號下游的菸鹼醯胺腺嘌呤二核苷酸(磷酸) [NAD(P)]對AtCAPE9誘導的氣孔免疫反應至關重要。核黃素同樣在促進局部和系統氣孔免疫反應中發揮了關鍵作用。綜合而言，我們的研究表明，NHP和riboflavin作為AtCAPE9的關鍵下游調節因子，調控了支撐SAR和氣孔免疫的代謝及免疫信號傳導。

Systemic acquired resistance (SAR) is a plant-wide defense response initiated by local pathogen infection, primarily mediated by salicylic acid (SA) and pathogenesis-related protein 1 (PR1). Our recent study identified AtCAPE9, a cytokine peptide derived from PR1, as a critical regulator of both local and systemic immune responses triggered by the bacterial flagellin-derived peptide flg22. While AtCAPE9 was shown to function as a key mediator in SA signaling, the metabolic pathways involved in its regulation of SAR remained unclear. To elucidate these pathways, we conducted a metabolomic analysis of Arabidopsis leaves and phloem exudates from the plants treated with AtCAPE9 using liquid chromatography-mass spectrometry (LC-MS/MS). This analysis revealed significant alterations in primary and secondary metabolites, with N-hydroxypipecolic acid (NHP) and riboflavin emerging as key metabolites elevated in both leaves and phloem exudates. Functional studies confirmed the essential role of NHP in AtCAPE9-induced immunity, as Arabidopsis mutants deficient in flavin-dependent monooxygenase 1 (FMO1), deficient in NHP biosynthesis, failed to exhibit AtCAPE9-triggered local and systemic immune responses. Furthermore, we found that nicotinamide adenine dinucleotide (phosphate) [NAD(P)], a known downstream component of NHP signaling, is required for AtCAPE9-induced stomatal immunity. Riboflavin also played a pivotal role in promoting both local and systemic stomatal immune responses. These findings demonstrate that NHP and riboflavin acts as an downstream of AtCAPE9, orchestrating metabolic and immune signaling pathways that underpin SAR and stomatal immunity in Arabidopsis.