Fig. 5

Expression of mutant UPF3B proteins affects neuronal differentiation of neural stem cells. Neural stem cells were transfected with pIRES-GFP (empty vector) or pIRES-GFP derivatives expressing the indicated UPF3B proteins. Differentiation was induced after 24 h, and analysed after 3 days and 6 days of differentiation by microscopy. Transfected cells were identified and visualised by GFP fluorescence. The analysis of neuron differentiation was performed as described in Methods. a. Representative pictures of neurons with tracing, and tracing of 2 additional neurons after 6 days of differentiation illustrating a reduction in branching in neurons expressing mutant UPF3B proteins. The scale bars represent 20 μm. Tracings are to scale with primary neurites traced with thicker lines then branches protruding from primary neurites. b, c. Branching was analysed after 3 days and 6 days as described and results are presented as box plots. The box plots show the median, with boxes spanning 25^th- 75^th percentiles and the crossbars indicating the 10^th and 90^th percentiles. Experiments were done independently 3 times with similar outcomes and the data were combined for presentation. Numbers of cells analysed are indicated. Asterisks indicate significantly reduced branching in neurons expressing mutant UPF3B proteins compared to neurons transfected with the empty vector (Kruskal Wallis test followed by Dunn's multiple comparison test; P < 0.05)