Inhibition of sonic hedgehog autoprocessing in cultured mammalian cells bysterol deprivation

Sonic hedgehog (Shh) is a signaling molecule that is important for defining patterning in the developing vertebrate central nervous system. After tran slation, Shh autoproteolyzes and covalently attaches cholesterol to the new ly formed carboxyl terminus, a modification crucial for normal Shh signalin g. Presented here is evidence that acute severe sterol deprivation in cultu red Chinese hamster ovary cells expressing mouse Shh (mShh) inhibits autopr ocessing of the protein. These conditions allowed the first detailed kineti c analysis of mShh autoprocessing and turnover rates revealing that cells r apidly degrade both precursor and mature mShh regardless of sterol content and sterol deprivation increases the rate of precursor degradation. Inhibit ion of mShh autoprocessing also allowed the determination of the subcellula r localization of mShh precursor which accumulates in a pre-medial Golgi in tracellular compartment. Finally, the precursor form of mShh that results f rom autoprocessing inhibition appears to accumulate as an amide rather than a stable thioester.