Figure 10.

mTOR regulates KIF1B expression and Mbp mRNA transport in differentiating oligodendrocytes in vitro. A, Western blot analysis showing decreased KIF1B expression with mTOR inhibition during oligodendrocyte differentiation in vitro. Rapamycin was added at the initiation of differentiation as for prior experiments (Fig. 1A); n = 3; p= 0.041 at D2, p = 0.007 at D3. B, Experimental paradigm of OPC differentiation plus or minus rapamycin addition following 2 d of normal differentiation. C, Western blot analysis of KIF1B expression after delayed rapamycin (R) treatment in vitro; n = 5, p = 0.006 at D4, p = 0.011 at D5. D, Representative images and percentage of MBP protein localization restricted to the cell body in rapamycin-treated oligodendrocytes at D4 in delayed addition paradigm; n = 4 Ctrl, n = 3 Rapa, p = 0.0008. E, Western blot analysis of MBP expression after delayed rapamycin treatment in vitro; n = 3, p = 0.007 at D3; p = 0.001 at D4; p = 0.032 at D5. F, G, Representative images of DIC and Mbp mRNA fluorescence (F) and quantification of Mbp mRNA localization (G) following single-molecule FISH for Mbp mRNA on primary rat OPCs after 7 d of differentiation in the absence (F, top panels; G control) or presence (F, bottom panels; G Rapa) of rapamycin from the start of differentiation. Scale bar, 5 µm. Mbp mRNA is reduced in the cellular processes of rapamycin-treated OPCs with a corresponding accumulation in the cytoplasm and nucleus; n = 51 Ctrl, n = 53 Rapa. *p < 0.001 for Control versus Rapa in all compartments.