Cellular and subcellular localization of the HVRP1 protein.

<p><b>A</b>) Immunohistochemical analysis of HVRP1 distribution in cerebellar tissue. Cerebellar cortical section from a wild-type rat stained with polyclonal antibody raised against HVRP1 (red). GC: granule cell layer, PC: Purkinje cell layer, ML: molecular layer, scale bar: 50 µm. <b>B</b>) Higher magnification of granule neurons expressing HVRP1, from (A). Scale bar is 10 µm. <b>C</b>) Ultrastructural localization of HVRP1 in the rat cerebellar GC layer visualized with the pre-embedding immunogold method. Positive labeling is in black, in the glomeruli, at the dendritic claws of granule cells surrounding mossy fiber terminals. Scale bar is 0.1 µm. <b>D</b>) Basic circuit diagram of the cerebellar cortex showing the location of HVRP1 in the dendrites and cell bodies of GCs. GO: Golgi cell, GR: glomerulus, MF: mossy fiber, PC: Purkinje cell. <b>E</b>) Confocal images of cultured mouse cerebellar granule neurons fixed and immunostained at 8 DIV. HVRP1 signal is shown in red (antibody dilution 1∶1000), DAPI signal is in blue. <b>F</b>) Staining as in (E) but with anti-HVRP1 antibody at dilution 1∶500 pre-incubated with HVRP1 antigen peptide (see Methods). Scale bars are 10 µm. <b>G</b>) Confocal image of live HEK293A cells expressing recombinant EGFP-tagged human HVRP1 (green) and labeled with the plasma membrane marker FM-464 (red). Scale bar is 10 µm. <b>H</b>) Western blot of total protein extracts from HEK293A cells transfected with hHVRP1 (H) and non transfected (N). AP indicates pre-incubation of anti-HVRP1 antibody with antigen peptide.</p>