Single-Cell Metabolite Profiling of Stalk and Glandular Cells of Intact Trichomes with Internal Electrode Capillary Pressure Probe Electrospray Ionization Mass Spectrometry

In
this report, we developed the pressure probe electrospray ionization-mass
spectrometry with internal electrode capillary (IEC-PPESI-MS) which
enables high spatial-resolution cell sampling, precise postsampling
manipulation, and high detection sensitivity. Using this technique,
a comparative in situ single-cell metabolite profiling
of stalk and glandular cells, the two adjacent cell types comprising
a trichome unit in tomato plants (Solanum lycopersicum L.), were performed to clarify the extent of metabolic differentiation
between two cell types as well as among different types of trichomes.
Owing to high sensitivity of the system, less than a picoliter cell
sap from a single stalk cell sufficiently yielded a number of peaks
of amino acids, organic acids, carbohydrates, and flavonoids. The
minimal cell sap removal from a stalk cell without severe disturbance
of trichome structure enabled sequential analysis of adjacent glandular
cell on the same trichome, which showed the presence of striking differences
in metabolite compositions between two adjacent cell types. Comparison
among different types of trichome also revealed significant variations
in metabolite profiles, particularly in flavonoids and acyl sugars
compositions. Some metabolites were found only in specific cell types
or particular trichome types. Although extensive metabolomics analysis
of glandular cells of tomato trichomes has been previously documented,
this is the first report describing cell-to-cell variations in metabolite
compositions of stalk and glandular cells as well as in different
trichome types. Further application of this technique may provide
new insights into distinct metabolism in plant cells displaying variations
in shape, size, function and physicochemical properties.

本研究中，我们开发了搭载内部电极毛细管的压力探针电喷雾电离质谱（internal electrode capillary-pressure probe electrospray ionization-mass spectrometry, IEC-PPESI-MS），该技术可实现高空间分辨率细胞采样、精准的采样后操作以及高检测灵敏度。利用该技术，我们针对番茄（Solanum lycopersicum L.）毛状体单元的两种相邻细胞类型——柄细胞与腺细胞，开展了原位单细胞代谢物谱的对比分析，以明确这两种细胞类型以及不同类型毛状体之间的代谢分化程度。得益于该系统的高灵敏度，单根柄细胞仅需不足1皮升的细胞液，即可检测到氨基酸、有机酸、碳水化合物及黄酮类化合物的多个特征峰。在不对毛状体结构造成严重破坏的前提下，仅微量移除柄细胞的细胞液，即可对同一毛状体上相邻的腺细胞进行连续分析，结果显示两种相邻细胞类型的代谢物组成存在显著差异。对不同类型毛状体的比较分析同样揭示出其代谢物谱存在显著差异，尤其是在黄酮类化合物与酰基糖的组成方面。部分代谢物仅在特定细胞类型或特定毛状体类型中被检出。尽管此前已有针对番茄毛状体腺细胞的大规模代谢组学研究，但本研究为首篇报道柄细胞与腺细胞之间以及不同毛状体类型之间细胞间代谢物组成差异的报告。该技术的进一步应用或将为形态、大小、功能及理化性质存在差异的植物细胞的特有代谢机制提供新的研究视角。