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. 2017 May 19;7:2149. doi: 10.1038/s41598-017-02202-4

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© The Author(s) 2017

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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TrkB receptors accumulate in EEA1 and Rab7 positive endosomes in EndophilinA triple knockouts. (A) HA-conjugated ProteinA beads (or IgG control beads) were used to pull down HA-TrkB (left) and EndophilinA1, A2, or A3-GFP (right) in co-transfected Hek293 cells in the presence or absence of 100 ng/ml BDNF. (B) Representative images of wild-type and EndophilinA TKO mouse neuronal cultures transfected with TrkB-GFP and the RFP-tagged endosomal markers EEA1 or Rab7. Arrows indicate TrkB colocalizing with EEA1 or Rab7. Scale bar = 10 µm. (C) Quantitation of colocalization of TrkB-GFP with RFP-tagged EEA1 or Rab7 (D), in wild-type and EndophilinA TKO neurons. In EndophilinA triple knockout neurons, more TrkB was found in EEA1 and Rab7 positive endosomes in both the presence and absence of BDNF compared to control; n = 30 images from 4 independent cultures, significance determined by unpaired two-tailed Student’s t-test compared to non-stimulated control and between non stimulated TKO and BDNF treated TKO, error = SEM; **p < 0.01, ***p < 0.001.