Comparative Analysis of Mixed-Transcriptomes between Tomato and Rice Infected with the Plant Pathogenic Fungi Botrytis cinerea and Magnaporthe oryzae

Abstract

真菌病原体持续发展不同的策略来规避植物防御，以便攻击并进入宿主。灰葡萄孢 (Botrytis cinerea) 是番茄的一种毁灭性坏死营养型病原体。然而，之前的大多数研究只探索了有限的感染阶段，并且病原体末端的反应仍然很大程度上未经调查。因此，本研究旨在分析番茄叶片五个感染阶段的高质量病原体富集混合转录组。还进行了番茄和水稻之间的直系同源分析，以确定当宿主植物感染真菌病原体时保守和非保守的植物-宿主反应。结果，在感染后23小时、40小时和47小时（hpi）观察到与番茄病原体反应相关的几个差异表达基因（DEG）；发现了 144 个 DEG，其中 2.6% 在三个时间点上持续表达。虽然番茄 DEG 编码抗病蛋白、受体样激酶以及与茉莉酸 (JA) 和乙烯 (ET) 介导的反应相关的发病机制相关蛋白，但也表达了一些毒力和生长相关基因以及候选效应子灰霉病菌。有趣的是，这种真菌表达了一些与氧化还原反应和解毒有关的防御相关基因。因此，表明病原体反应并非完全被动，而是表现出主动防御策略，可能涉及宿主防御反应的调节或抵消效应子的分泌。此外，比较转录组分析显示，84%的直系同源DEG在番茄和水稻之间高度保守，其中10.23%显示出与病原体防御反应相关的功能重要性。此外，利用 1DCNN 模型和引导反向传播进行顺式调控元件富集和深度学习分析，确定 WRKY 和 ERF 是与番茄生物胁迫响应表达模式相关的相关调控基序。此外，对番茄和水稻转录起始位点（TSS）的大规模比较分析揭示了有趣的模式。发现基因表达水平与GC-skew和AT-skew呈正相关，表明TSS周围的核苷酸组成可能对基因表达产生影响。虽然这两个物种都表现出保守的调控元件，例如上游 TSS 中的 TATA 盒，但在下游区域观察到了变化。具体来说，水稻显示出番茄中不存在的 Y 补丁信号。这些发现表明下游转录调控中有专门的系统和独特的调控元件参与，有助于植物基因组中 TSS 的多样性。

Fungal pathogens persistently develop different strategies to circumvent plant defenses so that they can attack and enter their host. Botrytis cinerea is a devastating necrotrophic pathogen of tomato. However, most previous studies have explored limited infection stages, and the response at the pathogen end still remains largely uninvestigated. Therefore, this study aimed to analyze high-quality pathogen-enriched mixed-transcriptome at five infection stages of tomato leaves. Ortholog analysis between tomato and rice was also carried out to identify the conserved and non-conserved plant-host responses when host plants were infected with a fungal pathogen. As a result, several differentially expressed genes (DEGs) related to pathogen response in tomato were observed at 23, 40, and 47 hours post-infection (hpi); 144 DEG’s were found, and 2.6% were constantly expressed across three time points. While tomato DEGs encoding disease-resistance proteins, receptor-like kinases, and pathogenesis-related proteins associated to jasmonic acid (JA) and ethylene (ET)-mediated responses, some virulence- and growth-related genes as well as candidate effectors were expressed in B. cinerea. Interestingly, the fungus expressed some defense-related genes for redox reaction and detoxification. Thus, it is suggested that the pathogen response is not completely passive, but rather exhibits an active defense strategy, potentially involving modulation of host defense responses or secretion of counteracting effectors. In addition, comparative transcriptome analysis revealed that 84% of the orthologous DEGs were highly conserved between tomato and rice, and 10.23% of which showed functional importance related to pathogen defense response. Moreover, cis-regulatory elements enrichment and deep learning analysis with the 1DCNN model and guided-backpropagation identified WRKY and ERF as relevant regulatory motifs associated with biotic stress-responsive expression patterns in tomato. Furthermore, the large-scale comparative analysis of transcription start sites (TSS) in tomato and rice revealed intriguing patterns. A positive correlation was found between gene expression levels and both GC-skew and AT-skew, indicating that the nucleotide composition around the TSS may exert influence on gene expression. While both species exhibited conserved regulatory elements, such as the TATA box in the upstream TSS, variations were observed in the downstream regions. Specifically, rice displayed Y-patch signals that were absent in tomato. These findings suggest the involvement of specialized systems and distinct regulatory elements in the downstream transcriptional regulation, contributing to the diversity of TSS in plant genomes.