Biological Sciences and Bioengineering Program, Sabanci University, Tuzla, Istanbul, Turkey.
Planta. 2012 Oct;236(4):1081-92. doi: 10.1007/s00425-012-1657-3. Epub 2012 May 9.
An autophagy-related gene Atg8 was cloned for the first time from wild emmer wheat, named as TdAtg8, and its role on autophagy under abiotic stress conditions was investigated. Examination of TdAtg8 expression patterns indicated that Atg8 expression was strongly upregulated under drought stress, especially in the roots when compared to leaves. LysoTracker(®) red marker, utilized to observe autophagosomes, revealed that autophagy is constitutively active in Triticum dicoccoides. Moreover, autophagy was determined to be induced in plants exposed to osmotic stress when compared to plants grown under normal conditions. Functional studies were executed in yeast to confirm that the TdATG8 protein is functional, and showed that the TdAtg8 gene complements the atg8∆::kan MX yeast mutant strain grown under nitrogen deficiency. For further functional analysis, TdATG8 protein was expressed in yeast and analyzed using Western immunoblotting. Atg8-silenced plants were exposed to drought stress and chlorophyll and malondialdehyde (MDA) content measurements demonstrated that Atg8 plays a key role on drought stress tolerance. In addition, Atg8-silenced plants exposed to osmotic stress were found to have decreased Atg8 expression level in comparison to controls. Hence, Atg8 is a positive regulator in osmotic and drought stress response.
首次从野生二粒小麦中克隆了一个与自噬相关的基因 Atg8，命名为 TdAtg8，并研究了其在非生物胁迫条件下对自噬的作用。检测 TdAtg8 的表达模式表明，干旱胁迫下 Atg8 的表达强烈上调，尤其是与叶片相比时，根部的表达上调更为明显。利用 LysoTracker(®) red 标记物观察自噬体，表明在 Triticum dicoccoides 中自噬是持续活跃的。此外，与正常条件下生长的植物相比，发现渗透胁迫诱导了植物的自噬。在酵母中进行的功能研究证实 TdATG8 蛋白是功能性的，并表明 TdAtg8 基因可补充在氮饥饿下生长的 atg8∆::kan MX 酵母突变体。为了进一步进行功能分析，在酵母中表达了 TdATG8 蛋白，并通过 Western 免疫印迹进行了分析。对 Atg8 沉默的植物进行干旱胁迫处理，并测量叶绿素和丙二醛（MDA）含量，结果表明 Atg8 在干旱胁迫耐受中起关键作用。此外，与对照相比，暴露于渗透胁迫下的 Atg8 沉默植物的 Atg8 表达水平降低。因此，Atg8 是渗透和干旱胁迫响应的正调控因子。
