PROTEOLYTIC PROCESSING YIELDS 2 SECRETED FORMS OF SONIC-HEDGEHOG

sonic hedgehog (Shh) is expressed in tissues with known signalling cap acities, such as the notochord, the floor plate of the central nervous system, and the zone of polarizing activity in the limb. Several line s of evidence indicate that Shh is involved in floor plate induction, somite patterning, and regulation of anterior-posterior polarity in th e vertebrate limb. In this report, we investigate the biochemical beha vior of Shh in a variety of expression systems and embryonic tissues. Expression of mouse Shh in Xenopus oocytes, COS cells, and baculovirus -infected insect cells demonstrates that in addition to signal peptide cleavage and N-linked glycosylation, chicken and mouse Shh proteins u ndergo additional proteolytic processing to yield two peptides with mo lecular masses of approximately 19 kDa (amino terminus) and 27 kDa (ca rboxy terminus), both of which are secreted. In transfected COS cells, we show that the 19-kDa peptide does not accumulate significantly in the medium unless heparin or suramin is added, suggesting that this pe ptide associates with the cell surface or extracellular matrix. This r etention appears to depend on sequences in the carboxy-terminal part o f the peptide. Finally, detection of the 19-kDa product in a variety o f mouse and chicken embryonic tissues demonstrates that the proteolyti c processing observed in cell culture is a normal aspect of Shh proces sing in embryonic development. These results raise the possibility tha t amino- and carboxyl-terminal regions of Shh may have distinct functi ons in regulating cell-cell interactions in the vertebrate embryo.