Extended Data Fig. 6: Localization and effects of cleaved EBP1 fragments in hippocampal neurons.
From: EBP1 potentiates amyloid β pathology by regulating γ-secretase
a, GFP-EBP1 fragments transfected hippocampal primary neurons at DIV9 were analyzed to show the nuclear translocation of 205–394 fragment. (left) After 24 h transfection, neurons were fixed with 4% PFA and stained with anti-NeuN antibody (purple). (right) Nuclei of GFP-plasmids overexpressed neurons were stained with DAPI. The expression pattern of each panel was depicted to the graphs. b, Hippocampal primary neurons at DIV9 were overexpressed the GFP-EBP1 fragments and stained with anti-Tuj1 antibody (red). White arrows indicate the axons of GFP-transfected neurons. Quantification of axonal length (μm) of GFP-positive neurons (right). The measured GFP-positive neurons were selected randomly and all data were normalized to GFP-Mock. Scale bar, 20 μm. n = 3. c, DIV9 5X-FAD primary hippocampal neurons were transfected with the indicated RFP-tagged plasmids and stained with anti-Tuj1 antibody (green). White arrows indicate the axons of GFP-transfected neurons. Axon length (μm) with Tuj1-positive neurons were quantified (right). The measured RFP-positive neurons were selected randomly and all data were normalized to RFP-Mock. Scale bar, 20 μm. n = 3. d, GFP-EBP1 plasmids were transfected to primary neurons at DIV9 and neurons were subjected to immunocytochemistry with anti-LAMP1 (red) and anti-EEA1 (blue) antibodies. White arrows mean the GFP-colocalization with EEA1-LAMP1 signals. Data are presented as mean values ± SEM (b, c). *p < 0.05, **p < 0.005, ***p < 0.001, ****p < 0.0001, ns; not significant (b, c). Student’s two-tailed unpaired t-test (b, c).
