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Figure 7. The tail domain of BMPRII is essential for the interaction with Trpc1, and noggin modulates the BMPRII-Trpc1 interaction. (A) Schematic illustration of the full-length BMPRII and the tail domain-deleted construct (BRII-∆TD). The yellow, black, grey and red rectangles show the signal peptide (SP); transmembrane domain (TM), kinase domain, and tail domain respectively. (B) Representative western blots showing the immunoprecipitation data acquired when analysing the protein-protein interaction between BMPRII and Trpc1. Embryos at the 4-cell stage were either injected (+) or not (−) with BMPRII-HA (200 pg/cell) or BRII-∆TD-HA (200 pg/cell), along with Myc-Trpc1 (200 pg/cell) into all the blastomeres. Animal caps were then prepared at stage 8–9, lysed and subjected to immunoprecipitation. ns, non-specific band. Western blot data were revealed by enhanced chemiluminescence (ECL; n = 4 independent experiments). (C) Representative western blots showing the interaction between BMPRII and TRPC1 in the presence of noggin. Animal caps were collected at stage 8–9 from embryos injected with BMPRII-HA (200 pg/cell) and Myc-Trpc1 (200 pg/cell) into all the blastomeres at the 4-cell stage, after which they were either incubated (+) or not (−) with 2 µg/mL noggin for 15 min, and then lysed and subjected to immunoprecipitation with anti-Myc antibody. The western blot data were revealed by ECL (n = 3 independent experiments). (D) Quantification of the BMPRII-HA fraction associated with Myc-Trpc1. Ratios of co-immunoprecipitated BMPRII-HA: Myc-Trpc1 were calculated in the absence or presence of noggin using the Bio-Rad ChemiDoc Image Lab software 5.2.1. There data represent the mean ± from 3 independent experiments. The asterisk indicates data that are significantly different at p < 0.05 when using the Mann-Whitney test. The full-length blots are presented in Supplementary figure S7. Image published in: Néant I et al. (2019) © The Author(s) 2019. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View

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