The cell wall integrity (CWI) signaling pathway regulates yeast cell wall biosynthesis, cell division, and responses to external stress. The cell wall, comprised of a dense network of chitin, β-1,3- and β-1,6- glucans, and mannoproteins, is very thin, <100 nm. Alterations in cell wall composition may activate the CWI pathway. Saccharomyces cerevisiae, a model yeast, was used to study the role of individual wall components in altering the structure and biophysical properties of the yeast cell wall. Near-field Fourier transform infrared spectroscopy (nano-FT-IR) was used for the first direct, spectrochemical identification of cell wall composition in a background (wild-type) strain and two deletion mutants from the yeast knock-out collection: kre6Δ and knr4Δ. Killer toxin resistant 6 (Kre6) is an integral membrane protein required for biosynthesis of β-1,6-glucan, while Knr4 is a cell signaling protein involved in the control of cell wall biosynthesis, in particular, biosynthesis and deposition of chitin. Complementary spectral data were obtained with far-field (FF)-FT-IR, in transmission, and with attenuated total reflectance (ATR) spectromicroscopy with 3 –10 μm wavelength-dependent spatial resolution. The FF-FT-IR spectra of cells and spectra of isolated cell wall components showed that components of the cell body dominated transmission spectra and were still evident in ATR spectra. In contrast, the nano-FT-IR at ∼25 nm spatial resolution could be used to characterize the yeast wall chemical structure. Our results show that the β-1,6-glucan content is decreased in kre6Δ, while all glucan content is decreased in the knr4Δ cell wall. The latter may be thinner than in wild type, since not only are mannan and chitin detectable by nano-FT-IR, but also lipid membranes and protein, indicative of cell interior.

中文翻译：

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纳米傅里叶变换红外光谱揭示了缺少 Knr4 和 Kre6 的酿酒酵母细胞壁重塑

细胞壁完整性 (CWI) 信号通路调节酵母细胞壁生物合成、细胞分裂和对外部应激的反应。细胞壁由几丁质、β-1,3-和β-1,6-葡聚糖以及甘露糖蛋白组成的致密网络组成，非常薄，<100 nm。细胞壁组成的改变可能激活 CWI 途径。酿酒酵母（Saccharomyces cerevisiae）是一种模型酵母，用于研究单个壁成分在改变酵母细胞壁的结构和生物物理特性中的作用。近场傅立叶变换红外光谱 (nano-FT-IR) 用于首次直接光谱化学鉴定背景（野生型）菌株和酵母敲除集合中的两个缺失突变体的细胞壁组成：kre6Δ 和knr4Δ。杀伤毒素抗性 6 (Kre6) 是 β-1,6-葡聚糖生物合成所需的整合膜蛋白，而 Knr4 是一种细胞信号蛋白，参与细胞壁生物合成的控制，特别是几丁质的生物合成和沉积。补充光谱数据通过远场 (FF)-FT-IR、透射和衰减全反射 (ATR) 光谱显微镜获得，空间分辨率为 3 –10 μm 波长相关。细胞的 FF-FT-IR 光谱和分离的细胞壁成分的光谱表明，细胞体成分主导透射光谱，并且在 ATR 光谱中仍然很明显。相比之下，~25 nm 空间分辨率的纳米 FT-IR 可用于表征酵母壁的化学结构。我们的结果表明，kre6Δ 中的 β-1,6-葡聚糖含量降低，而 knr4Δ 细胞壁中的所有葡聚糖含量均降低。后者可能比野生型更薄，因为纳米 FT-IR 不仅可以检测甘露聚糖和几丁质，还可以检测脂质膜和蛋白质，从而指示细胞内部。