使用受激发射损耗显微镜对酵母过氧化物酶体膜蛋白及其结合伴侣进行共定位的实验方案。

Protocol for the colocalization of yeast peroxisomal membrane proteins and their binding partners using stimulated emission depletion microscopy.

作者信息

Mol Frank N, de Lange Eline M F, van der Klei Ida J, Vlijm Rifka

机构信息

Molecular Biophysics, Zernike Institute for Advanced Materials, University of Groningen, Nijenborgh 7, 9747 AG Groningen, the Netherlands.

Molecular Biophysics, Zernike Institute for Advanced Materials, University of Groningen, Nijenborgh 7, 9747 AG Groningen, the Netherlands; Molecular Cell Biology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 7, 9747 AG Groningen, the Netherlands.

出版信息

STAR Protoc. 2025 Aug 22;6(3):103998. doi: 10.1016/j.xpro.2025.103998.

DOI:10.1016/j.xpro.2025.103998

PMID:40848258

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12397933/

Abstract

Peroxisomes are highly dynamic organelles that play crucial roles in cellular metabolism. Here, we present a protocol to investigate peroxisomal membrane proteins and their binding partners in the yeast Hansenula polymorpha using stimulated emission depletion (STED) nanoscopy. We describe steps for strain construction to facilitate specific STED-compatible labeling, automated STED imaging, and semi-automated data analysis. This protocol enables examination of the dynamic alterations in the colocalization of the peroxisomal membrane protein Pex3 and its binding partner Atg30. For complete details on the use and execution of this protocol, please refer to de Lange et al..

摘要

过氧化物酶体是高度动态的细胞器，在细胞代谢中发挥着关键作用。在此，我们展示了一种使用受激发射损耗（STED）纳米显微镜研究多形汉逊酵母中过氧化物酶体膜蛋白及其结合伴侣的方法。我们描述了构建菌株以促进特定的与STED兼容的标记、自动化STED成像和半自动数据分析的步骤。该方法能够检测过氧化物酶体膜蛋白Pex3及其结合伴侣Atg30共定位的动态变化。有关此方法的使用和执行的完整详细信息，请参考德朗热等人的研究。