Specific subdomain localization of ER resident proteins and membrane contact sites resolved by electron microscopy
2021
The endoplasmic reticulum (ER) is a large, single-copy, membrane-bound organelle that comprises an elaborate 3D network of diverse structural subdomains, including highly curved tubules, flat sheets, and parts that form contacts with nearly every other organelle. The dynamic and complex organization of the ER poses a major challenge on understanding how its functioning - maintenance of the structure, distribution of its functions and communication with other organelles - is orchestrated. In this study, we resolved a unique localization profile within the ER network for several resident ER proteins representing a broad range of functions associated with the ER using immuno-electron microscopy and calculation of a relative labeling index (RLI). Our results demonstrated the effect of changing cellular environment on protein localization and highlighted the importance of correct protein expression level when analyzing its localization at subdomain resolution. We present new software tools for anonymization of images for blind analysis and for quantitative assessment of membrane contact sites (MCSs) from thin section transmission electron microscopy micrographs. The analysis of ER-mitochondria contacts suggested the presence of at least three different types of MCSs that responded differently to changes in cellular lipid loading status.
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