Immunocytochemistry confirms increased nuclear expression of Ube1 in Wldsmouse cerebellum. Confocal micrographs of cerebellar granule cells from (a-c) Wldsand (d-f) wild-type mice. Ubiquitin-activating enzyme E1 (Ube1) is shown in green and the nuclear marker TOPRO3 is shown in blue (panels a and d show Ube1; panels b and e show TOPRO3; and panels c and f show both markers). Note how Ube1 protein appears to be more strongly expressed in the nuclei of Wldscerebellar neurons, whereas TOPRO3 and cytoplasmic levels of Ube1 appear unchanged. (g-i) Scatter plots (line indicates mean) of fluorescence intensity (see Materials and methods) of nuclear Ube1 (panel g), nuclear TOPRO3 (panel h), and cytoplasmic Ube1 (panel i). Only nuclear Ube1 was significantly increased in intensity in Wldsneurons (P < 0.001; by unpaired, two-tailed t-test). Scale bar 20 μm.