UC San Diego
Visualization of membrane-less granules in yeast and mammalian cells using modified fluorescence in-situ hybridization
- Author(s): Kopalle, Hema
- Advisor(s): Zid, Brian M
- et al.
Processing bodies (p-bodies) are transient, membrane-less constructs consisting of mRNA and RNA-binding proteins. Though constitutively expressed in mammalian cells, formation of these structures in yeast is induced by metabolic stress. While not all mRNAs localize to membrane-less granules, identifying their contents has proved difficult because of the structural nature of these droplets. It therefore becomes necessary to stabilize these granules to qualitatively observe mRNA localization. Here we present a new methodology for granule stabilization using a combination of UV cross-linking and the small dialdehyde Glyoxal. We also provide evidence suggesting Glyoxal to be an effective fixative for membrane less granule fixating in mammalian cells.