Expression analysis of the salt stress response in Arabidopsis mutants with defects in hair patterning. Arabidopsis thaliana

We performed an expression analysis of the response of seedling root tips to 1 hour of treatment with 140mM NaCl using mutants defective in root hair patterning. Cells are amazingly adept at integrating both external and internal cues to regulate transcriptional states. While internal processes such as differentiation and cell-type specification are generally understood to have an important impact on gene expression, very little is known about how cells utilize these developmental cues to regulate responses to external stimuli. Here we use the response to a well characterized environmental stress, high salinity, to obtain a global view of the role that cell identity plays in guiding transcriptional responses in the root of Arabidopsis. Our analysis is based on three microarray data sets we have generated that explore transcriptional changes spatially among 6 cell layers and 4 longitudinal regions or temporally along 5 time points after salt treatment. We show that the majority of the response to salt stress is cell-type specific resulting in the differential regulation of unique biological functions in subsets of cell layers. To understand the regulatory mechanisms controlling these responses we have analyzed cis-element enrichment in the promoters of salt responsive genes and demonstrate that known stress regulatory elements likely control responses to salt occurring in multiple cell types. Despite the extensive shift in transcriptional state that salt stress elicits, we are able to identify several biological processes that consistently define each cell layer and find that transcriptional regulators of cell-identity tend to exhibit robust cell-type specific expression. Finally, using mutants that disrupt cell-type specification in the epidermis, we reveal cell autonomous and non-autonomous effects when cell identity is altered. Together, these data elucidate a novel intersection between physiology and development and expand our understanding of how transcriptional states are regulated in a multi-cellular context. Keywords: Response analysis of mutants Overall design: Seedlings were grown for 5 days before being transferred to standard media or media supplemented with 140mM NaCl. One hour after transfer, a razor blade was used to cut the root ~5 mm above the root tip. Approximately 30 roots were pooled per replicate. Two biological replicates were performed per genotype, per condition.

本研究利用根毛发育模式缺陷的突变体，对140mM氯化钠（NaCl）处理1小时后的幼苗根尖响应开展了表达谱分析。
细胞具备卓越的信号整合能力，可同时感知外界与内部信号以调控转录状态。
尽管学界普遍认为细胞分化与细胞类型特化等内部过程对基因表达具有重要调控作用，但目前对于细胞如何利用这些发育信号调控对外界刺激的响应，仍知之甚少。
本研究以已被充分表征的环境胁迫——高盐胁迫的响应为模型，旨在全面解析细胞身份在拟南芥（Arabidopsis）根部转录响应调控中发挥的核心作用。
本分析基于我们生成的三套微阵列（microarray）数据集，分别从空间维度（覆盖6个细胞层与4个纵向区域）以及时间维度（盐处理后5个时间点）探究转录组的动态变化。
我们发现，大多数盐胁迫响应具有细胞类型特异性，使得不同细胞层亚群中独特的生物学功能出现差异调控。
为解析调控这些响应的分子机制，我们分析了盐响应基因启动子区域的顺式作用元件（cis-element）富集情况，证实已知的胁迫调控元件可能调控多类细胞中发生的盐响应过程。
尽管盐胁迫会引发转录状态的广泛重塑，我们仍鉴定出了稳定定义各细胞层的若干核心生物学过程，并发现细胞身份的转录调控因子往往呈现显著的细胞类型特异性表达模式。
最后，我们利用破坏表皮细胞类型特化的突变体，揭示了细胞身份改变时的细胞自主性（cell autonomous）与非自主性效应。
综上，本研究阐明了生理学与发育生物学之间的全新交叉领域，并加深了我们对多细胞环境中转录状态调控机制的理解。

关键词：突变体响应分析

实验设计概要：将幼苗培养5天后，分别转移至标准培养基或添加140mM氯化钠（NaCl）的培养基中。处理1小时后，使用手术刀片在根尖上方约5mm处切割根系。每个生物学重复混合约30条根系；每个基因型、每个处理条件均设置2个生物学重复。