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. 2020 Jul 13;28(1):123–138. doi: 10.1038/s41418-020-0587-z

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — a Localization of a cMyc-SEPN1 in murine primary myotubes. Skeletal muscle cells were transfected with a plasmid expressing a cMyc-SEPN1 protein, which co-localized with calnexin (an ER marker), with the outer mitochondrial membrane protein TOM40 and with VDAC2 (a MAM marker). Scale bar is 10 µm. b Protein components of subcellular fractions prepared from muscle (gastrocnemius) and cells (HeLa) revealed by western blot analysis. SEPN1 presence was shown using a specific antibody. IP3R3 and PDI were used as ER markers, β-tubulin as a cytosolic marker, Sigma 1-R as a MAM marker, cytochrome C (Cyt c) as a mitochondrial marker. H homogenate, Mp pure mitochondria, ER endoplasmic reticulum, MAM mitochondria-associated membranes, C cytosol. c Co-localization of SEPN1-Flag (red) and Sigma 1-R-EGFP (a MAM marker, green) in C2C12 (right panel) and HeLa cells (left panel). The lower-right panel displays the merged image of the two stains. The lower-left panels display the SEPN1 signal overlaid with MAMs (MAM boundaries are highlighted in gray) in a rainbow lookup table (LUT). MAMs-SEPN1: Manders coefficient for SEPN1 staining was calculated as the proportion of SEPN1 signal overlapping with the Sigma 1-R marker.