Journal: Proceedings of the National Academy of Sciences of the United States of America
Article Title: nArgBP2 regulates excitatory synapse formation by controlling dendritic spine morphology
doi: 10.1073/pnas.1600944113
Figure Lengend Snippet: nArgBP2 KD increases WAVE1/PAK/cofilin phosphorylation, and counteracting this cascade rescues nArgBP2 KD effects on spines. Cortical neurons infected with AAV-scrambled or shRNA-nArgBP2 at DIV6, lysed at DIV21, and immunoblotted with anti-WAVE1 and phospho-WAVE1 antibody (A), PAK and phospho-PAK antibody (B), and anticofilin and phosphocofilin antibody (C) are shown. IB, immunoblot. (D) Representative images of spine from dendrites of neurons expressing each set of constructs. Hippocampal neurons were transfected at DIV9 and fixed at DIV16. Control, scrambled; shRNA (sh), shRNA-nArgBP2; S3A, phosphodeficient mutant of cofilin; mSH3-1/2, first and second SH3 domains of nArgBP2 targeted to mitochondria; mSH3-3, third SH3 domain of nArgBP2 targeted mitochondria. (Lower) Enlarged images of the insets enclosed with rectangles. (Scale bars: Upper, 10 μm; Lower, 2 μm.) The numbers of mushroom-shaped spines (E and G) and filopodia (F and H) from each experimental group are shown. *P < 0.05 by ANOVA, followed by Tukey’s honest significant difference (HSD) post hoc test [n = 45 neurons from seven independent cultures (E and F) and n = 39 neurons from four independent cultures (G and H)].
Article Snippet: Rabbit polyclonal anticofilin antibody and phosphocofilin antibody were from Cell Signaling Technology.
Techniques: Infection, shRNA, Western Blot, Expressing, Construct, Transfection, Mutagenesis