Figure 1.

Subcellular distribution of recombinant Mover variants in cultured hippocampal neurons. (A) Schematic depicting the proteins tested. The numbers indicate amino acids in the primary structure of rat Mover. The black bar indicates the location of the Calmodulin (CaM) binding region. All proteins are tagged with monomeric EGFP (mGFP). (B–G) Inverted gray level epifluorescence images representing the localization of GFP-tagged rat Mover constructs in DIV14 rat hippocampal neurons. The square panels on the right represent zooms of the boxes. Mover-mGFP and 52-266-mGFP are known to accumulate at synapses and produce the corresponding punctate staining pattern. All other variants are homogeneously distributed throughout the cytoplasm, indicating that they fail to accumulate at synapses. A 20× objective was used for the overviews on the left, in order to capture an entire neuron. The exposure time was set to visualize fluorescence in the neurites. Using these setting, nuclear and somatic fluorescence is saturated, because these compartments have much bigger volume than neurites. The distribution of each construct was determined in more than 30 neurons on a total of six coverslips from three independent experiments.