Subcellular Localization of ESCRT-II in the Nematode C. elegans by Correlative Light Electron Microscopy

Abstract

In this chapter, we report a protocol to perform correlative light electron microscopy (CLEM) on adult Caenorhabditis elegans. We use a specific fixation protocol, which preserves both the GFP fluorescence and the structural integrity of the samples. Thin sections are first analyzed by light microscopy to detect GFP-tagged proteins and, subsequently, with transmission electron microscopy (TEM) to characterize the ultrastructural anatomy of cells. The superimposition of light and electron images allows determining the subcellular localization of the fluorescent protein. We used CLEM to characterize the subcellular localization of the C. elegans ESCRT-II component VPS-36. VPS-36 protein localization in C. elegans muscle cell is strongly correlated with the sarcoplasmic reticulum network. Together with genetic evidences, the CLEM data support a role for ESCRT-II proteins in sarcoplasmic reticulum membrane shaping.