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. 2020 Mar 20;10:5115. doi: 10.1038/s41598-020-61970-8

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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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Proteolysis of OLM and OS proteins is increased in Sfrp1^−/− retinas leading to an abnormal protein distribution. a, b) Frontal cryostat sections from 3 months-old wt retinas immunostained with secondary antibodies only (a) or with anti-ADAM10 (b). Sections are counterstained with Hoechst. Note the specific immune signal in the OLM (arrows) and in the INL where Müller glial cells are located (yellow arrows). (c–e) Western blot analysis of N-cadherin, PCDH21 and CRB1 content in total protein extracts from three weeks-old wt, Sfrp1^−/− and rd10 retinas, as indicated in the panels. For N-cadherin and PCDH21, membranes were probed with antibodies recognizing the respective C-terminal fragments (CTF). Graphs at the bottom of each panel indicate the rate of CTF proteolysis for N-cadherin (c) and PCDH21 (d), calculated as CTF/FL, or the CRB1 levels (e). Plotted values are normalized to α-tubulin. (f–k) Frontal cryostat sections from 6 months-old wt and Sfrp1^−/− retinas immunostained with antibodies against N-cadherin, β-catenin, and GS as indicated in the panels. Sections are counterstained with Hoechst. White arrows in g, k indicate the gaps in the OLM. Scale bar calibration in k applies to all panels. Abbreviations: CTF: C-terminal fragment; FL: full length; OLM: outer limiting membrane; ONL: outer nuclear layer.