Figure 6.

Detection of the Skt protein. (A) Western blot analysis to detect Skt protein using extracts from untreated BMT10 cells (lane 1), BMT10 cells transfected with vector (lane 2), BMT10 cells transfected with the Skt expression vector (lane 3), and extracts of the nucleus pulposus of caudal IVDs from 8-week-old wild-type mice (lane 4), Skt^Gt/+ mice (lane 5), and Skt^Gt/Gt mice (lane 6). An ∼150-kDa protein corresponding to the predicted molecular weight of 147 kDa was detected in lanes 3, 4, and 5, but not in lane 6. The amount of Skt protein was reduced in the Skt^Gt/+ mutant (lane 5) and was below the detectable level in Skt^Gt/Gt (lane 6). (B) Immunohistochemistry of frontal sections of the nucleus pulposus in upper caudal IVDs from adult 8-week-old mice using purified anti-Skt antibodies. Skt protein was detected in the cytoplasm of nucleus pulposus cells in wild-type (a and c), but not Skt^Gt/Gt, mice (b and d). (c and d) Higher magnification of the area indicated by the boxes in a and b, respectively. Bars, 200 μm.