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. 2019 Apr 15;9:6101. doi: 10.1038/s41598-019-42571-6

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© The Author(s) 2019

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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The sheet morphology of melanosomal amyloid essentially depends on functional O-glycosylation. Mel220 transfectants expressing human PMEL were treated for four days with the O-glycosylation inhibitor Benzyl-2-acetamido-2-deoxy-α-D-galactopyranoside or with the solvent DMSO alone. (A–C) Western blot analysis of SDS-lysed total membranes using PMEL-specific antibodies EPR4864 (PMEL C-term.) (A), HMB45 (sialylated RPT domain) (B, two different exposures of the same blot are shown), and 6777 (CAF) (C). (D,E) Quantitative EM analysis showing the number of fibril-containing organelles per cell [N = 15]. An unpaired two-tailed t-test was used to determine whether means are statistically different from the DMSO-treated sample (NS, not significant) (D). Representative electron micrographs are depicted (E). Quantification of the number of fibril-containing organelles displaying the collapsed amyloid sheet phenotype. Shown is the average from three independent experiments. An unpaired two-tailed t-test was used to determine whether means are statistically different from the DMSO-treated sample (***p < 0.0001) (E, lower panel).