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. 2007 Oct 17;27(42):11366–11375. doi: 10.1523/JNEUROSCI.2326-07.2007

Figure 6.

Figure 6.

Myosin Va controls localization of VAMP2 to distal processes of oligodendrocytes. A, Purified oligodendrocytes at the A2B5 stage of maturation were stained for myosin Va, VAMP2, or BERP. As A2B5 type oligodendrocytes elaborate processes, VAMP2 becomes more distally localized to process tips and filopodia. VAMP2 localized to lamellas and filopodia does not colocalize with myosin Va, whereas more proximal VAMP2, including in distal process, does colocalize with myosin Va. BERP and myosin Va colocalize in A2B5 type oligodendrocytes. The arrows indicate examples of lamellas. B, GFP-tagged DN-myosin Va was nucleofected into oligodendrocytes, which were stained for VAMP2. VAMP2 does not localize to filopodia of oligodendrocyte distal processes and instead forms collections in the distal processes that colocalize with GFP-myosin Va tail signal. The arrows show regions of colocalization between GFP and endogenous VAMP2. C, VAMP2 localization is disrupted by blocking antibodies to myosin Va but not myosin Vb. Purified oligodendrocytes were replated and transfected with myosin Va blocking antibodies or control rabbit antibodies (top figures) or with myosin Vb blocking antibodies or control mouse antibodies (bottom figures). Cells were cultured for 6 h and stained for VAMP2 (green), actin (red), and DAPI (blue). In most distal processes transfected with control or myosin Vb antibodies, VAMP2 localization to distal processes clearly occurs. Cells transfected with myosin Va blocking antibodies had significantly less VAMP2 staining. Data from myosin Va (squares) or control (diamonds) transfected cells is displayed in top graph. Data from myosin Vb (squares) or control (diamonds) transfected cells is displayed in the bottom graph. D, Similar to VAMP2, BERP staining intensity increased proximally after myosin Va blocking antibody transfection. However, there were no differences distally. Data from BERP (squares) or control (diamonds) transfected cells is displayed in the graph to the right. For both C and D graphs, staining intensity was converted to percentage intensity found at the proximal root of each process. Intensities were plotted per decile of process length to allow comparison of processes of differing length. Error bars represent SEM. The asterisk indicates p < 0.05 by t test. Scale bars, 10 μm.