Huang Jinliang, Wang Geng
Ministry of Education Key Laboratory of Bioinformatics, Cell Biology and Development Center, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Bio Protoc. 2019 Jun 5;9(11):e3255. doi: 10.21769/BioProtoc.3255.
Cytosolic rRNAs are highly dynamic and can be degraded under conditions such as apoptosis, starvation and magnesium depletion. The degradation is also related to their specific localization, as fractions of cytosolic ribosomes are localized on the surfaces of intracellular organelles, such as endoplasmic reticulum (ER) and mitochondria. Such localized translation facilitates translocation of nascent proteins into these organelles co-translationally, contributing to fast responses to cellular stresses and precise regulations of the organelle. Here, we describe a protocol to establish the system to investigate rRNA degradation on mitochondrial outer membrane or ER. The protocol consists of organelle isolation, rRNA degradation on organelles and agarose gel electrophoresis to examine the remaining rRNAs.
胞质核糖体RNA高度动态变化，在诸如细胞凋亡、饥饿和镁离子耗竭等条件下会发生降解。这种降解还与其特定定位有关，因为部分胞质核糖体定位于细胞内细胞器表面，如内质网（ER）和线粒体。这种局部翻译有助于新生蛋白质共翻译转运到这些细胞器中，从而对细胞应激做出快速反应并对细胞器进行精确调控。在此，我们描述了一种建立用于研究线粒体外膜或内质网上核糖体RNA降解系统的方案。该方案包括细胞器分离、细胞器上的核糖体RNA降解以及用于检测剩余核糖体RNA的琼脂糖凝胶电泳。
