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. 2021 Apr 14;6:148. doi: 10.1038/s41392-021-00535-1

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© The Author(s) 2021

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. 6 — GOLM1 interacts with N2ICD and modulates its downstream signaling. a Mass spectrometry (MS) analysis of GOLM1-associated proteins. Total cell lysates from DDK-GOLM1 expressed cells were subjected to affinity purification. The purified protein complex was resolved on SDS-PAGE and silver stained; then the bands were retrieved and analyzed by MS. b A diagram depicts GOLM1 interactors as detected by MS (Detailed information in Supplementary Information). c The interaction between GOLM1 and NOTCH2/N2ICD in 293T cells stably expressing DDK GOLM1 was detected by immunoprecipitation. d The interaction between GOLM1 and NOTCH2 (FL)/N2ICD in Caco-2 cells was detected by immunoprecipitation. e Diagrammatic representation of GOLM1 and its truncated forms. Based on sequence and structure analyses, cytoplasmic domain, transmembrane domain, and Golgi lumen domain are indicated. f Various GOLM1 truncation constructs tagged with DDK were co-transfected with GFP-NOTCH2 in 293T cells for domain mapping. Immunoprecipitation analysis was performed with anti-GFP or anti-DDK antibodies. g Diagrammatic representation of N2ICD and its truncated forms. h Various N2ICD truncation constructs tagged with GFP were co-transfected with DDK-GOLM1 in 293T cells for domain mapping. Immunoprecipitation analysis was performed with anti-GFP or anti-DDK antibodies. i Cellular fractionations from GOLM1-deficient Caco-2 cells transfected with various GOLM1 truncation constructs and control cells were analyzed by immunoblotting with the indicated antibodies. j The relative mRNA expression levels of Notch signaling downstream genes from GOLM1-deficient Caco-2 cells transfected with various GOLM1 truncation constructs and control cells were determined by qRT-PCR (the data are represented as the means ± SEM; **P < 0.01, ***P < 0.001; unpaired, two-tailed Student’s t test)