ABSTRACT

Autophagy is an important cytoprotective process that mediates degradation of dysfunctional or unnecessary cellular components. In the process of autophagy, a double-membrane organelle termed the autophagosome is formed to sequestrate portions of cytoplasm and subsequently delivered into lysosome or vacuole for degradation. The accumulation of autophagic bodies in the vacuoles after treatment with concanamycin A (ConcA) is a widely used protocol for monitoring the occurrence of autophagy in plants. Here, it was found that the cytoplasmic soluble GFP was accumulated in vacuoles upon ConcA treatment. Importantly, the GFP signal showed good colocalization with the autophagic marker mCherry-ATG8f in vacuoles based on two commonly used methods, the Pearson-Spearman correlation colocalization analysis and the plot profile analysis. Further results showed that the free GFP did not interact with ATG8s. Thus, analysis of accumulation and colocalization only in vacuoles is not a trustworthy way to judge whether degradation of cytoplasmic protein is dependent on the selective autophagy pathway in plants. In this short perspective, we propose several primary steps to distinguish that the cytoplasmic proteins are degraded by selective or bulk autophagy, hoping they could contribute to identify and clarify the selective autophagic cargos and receptors in plants.

摘要

自噬是一种重要的细胞保护过程，它介导功能失调或不必要的细胞成分的降解。在自噬过程中，形成称为自噬体的双膜细胞器以隔离部分细胞质并随后递送到溶酶体或液泡中进行降解。用康卡那霉素 A (ConcA) 处理后液泡中自噬体的积累是一种广泛用于监测植物自噬发生的方案。在这里，发现细胞质可溶性 GFP 在 ConcA 处理后在液泡中积累。重要的是，基于 Pearson-Spearman 相关共定位分析和图谱分析这两种常用方法，GFP 信号与液泡中的自噬标记 mCherry-ATG8f 显示出良好的共定位。进一步的结果表明，游离的 GFP 不与 ATG8 相互作用。因此，仅分析液泡中的积累和共定位并不是判断细胞质蛋白降解是否依赖于植物中的选择性自噬途径的可靠方法。在这个简短的观点中，我们提出了几个主要步骤来区分细胞质蛋白被选择性或大量自噬降解，希望它们有助于识别和阐明植物中的选择性自噬货物和受体。