Membrane depolarisation induced dissocation of PTEN-DBN and increases the phosphorylation of pS647-DBN.

<p>(A) Neurons were transfected at 7 DIV with mCherry-DBN and GFP-PTEN and incubated until 10 DIV, before treatment with KCl (20 mM) and FLIM measurements. Bar graphs represent the average FRET efficiency of 22 cells over 3 independent experiments + sem. *p<0.001. Scale bar: 10 µm (B) Cortical neurons from floxed PTEN mice expressing RFP or RFP-Cre were cultured for 10 DIV and treated with KCl (20 mM). (C) 11 DIV rat hippocampal neurons were treated with KCl (20 mM) for 3 minutes in the presence or absence of Wortmannin (wm) at 100 nM. Bar graph shows the relative band density of pS647-DBN in three independent experiments +sem; *p<0.05. (D) Schematic diagram illustrating signaling relationships. DBN phosphorylation at S647 is suppressed through physical interactions with PTEN. Synaptic activity induces a transient dissociation of the PTEN:DBN complex and allows maximal DBN-S647 phosphorylation to occur.