Fig. 7. Shh protein is processed in Disp mutant embryos. Western blot of lysate from wild-type, Disp E8^+/-, Disp E8^-/-, Shh^-/- embryos collected at 9.5 dpc and COS7 cells transfected with expression constructs that encode either the full-length Shh protein (Shh) or the unmodified N-terminal fragment (Shh-N) probed with anti-Shh antibodies. Approximately equal amounts of proteins were loaded onto each lane. Both unprocessed (Shh,.upper arrow) and processed (Shh-Np, lower arrow) forms of Shh are detected from COS7 cells expressing the full-length Shh and are absent in lysate from Shh mutant embryos. A major band running at the same position as processed Shh was detected in lysate from wild-type, Disp E8^+/- and Disp E8^-/- embryos. The doublet observed in COS7 cells transfected with Shh-N could represent Shh-N proteins with different lipid modifications at its N terminus. A nonspecific band (or immunoreactivity with another Hh protein) was detected in lysates from embryos only and conveniently serves as a loading control. A very faint band representing the unprocessed Shh can be detected in lysates from wild-type, Disp E8^+/- and Disp E8^-/- embryos upon longer exposure (data not shown).

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