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14-3-3θ is a Binding Partner of Rat Eag1 Potassium Channels

Figure 5

Endogenous expression of 14-3-3θ and rEag1 in neurons.

(A) Co-immunoprecipitation of 14-3-3θ and rEag1. Detergent solubilized proteins from the lysates of rat forebrain were immunoprecipitated (IP) with the anti-14-3-3θ (upper panel) or the anti-rEag1 antibody (lower panel), followed by immunoblotting (WB) analyses with the anti-14-3-3θ or the anti-rEag1 antibody. The non-immune mouse or rabbit IgG was used in parallel as negative control. Input volumes correspond to 5% of the total cell lysates used for immunoprecipitation. The arrowhead and arrow refers to the protein bands of 14-3-3θ and rEag1, respectively. (B) Immunofluorescence staining of rEag1 (left panels) and 14-3-3θ (middle panels) in cultured hippocampal neurons. The area highlighted in the white boxes is viewed under a higher magnification (I, II). Arrows label the sites of co-localization of 14-3-3θ and rEag1 (right panels), which displayed significant punctuate patterns over a wide region along the neurites. Scale bar, 25 µm. These co-immunoprecipitation and immunofluorescence data are representative of four to seven independent experiments.

Figure 5

doi: https://doi.org/10.1371/journal.pone.0041203.g005