Fig 1: CaMKv regulates spine morphogenesis via RhoA inhibition.(a–c) CaMKv knockdown dramatically increased RhoA activity but not Rac1 or Cdc42. (d) The RhoA-GEF Lfc, but not the other GEFs TIAM1 or Ephexin1, was pulled down from the synaptosome by FLAG-tagged CaMKv. (e) CaMKv was co-immunoprecipitated with Lfc in brain homogenate and synaptosome fractions. Immunoprecipitation by IgG served as the negative control. (f) FLAG-tagged Lfc and HA-tagged RhoA were co-expressed in HEK293T cells, and the lysate was subsequently incubated with recombinant GST-tagged CaMKv CaM-binding domain (CaM) or the CaMKv PEST domain. Lfc–RhoA interaction was specifically disrupted by the CaMKv CaM-binding domain but not the PEST domain. (g,h) Hippocampal neurons transfected with CaMKv shRNA or Scr were treated with the ROCK inhibitor Y27632 (10 μM) or vehicle (Veh) at 17 DIV for 6 h. Y27632 abolished the spine loss induced by CaMKv shRNA (scale bar, 10 μm; n=30 dendrites per condition, ***P<0.001, one-way analysis of variance (ANOVA), Tukey's multiple comparison test). (i,j) Cultured hippocampal neurons were treated with the AMPA receptor blocker NBQX (20 μM) from 9 to 16 DIV. NBQX reduced CaMKv expression (n=3 independent experiments, *P<0.05, Student's t-test). (k,l) Hippocampal neurons transfected at 7 DIV with the indicated constructs were treated with NBQX (20 μM) or vehicle (Veh) from 9 to 16 DIV. The spine loss on long-term AMPA receptor blockade was rescued by CaMKv overexpression (scale bar, 10 μm; n=30 dendrites per condition, ***P<0.001, one-way ANOVA, Tukey's multiple comparison test).