Ma-Derived Neurite Promoting with Protease Inhibitory Activity Belongs to the Protease Nexins-

Sergio Gioor: Karel Odink,t Joachim Guenther;“* anspeter Nick: and Denis Monard” * Friedrich Miescher lnstitut CH-4002 Basei, Switzerland r Biotechnology Department Ciba-Geigy Ltd Basel, Switzerland giia-derived neurite promoting factor (GdNPF) has serine protease inhibitory activity and in addition regulates the migration of neuronal cells. cDNA clon- ing of GdNpF is necessary for studying the physiologi- cal relevance and the mode of action of this protein and similar cell-derived protease inhibitors. Xenopus oocytes injected with rat glioma cells mRNA release this inhibitor. A rat cDNA clone coding for the previ- ously purified glia-derived neurite promoting factor (GdNPF) was isolated upon hybridization-selected translation, followed by immunoprecipitation. The cor- rect identity of this cDNA is proven by the presence sequence coding for a tryptic fragment from pure PF. Northern analysis indicates that GdNPF mRNA is found almost exclusively in brain tissue and could be developmentally regulated. The same cDNA clone has been used to isolate full-length rat and hu- man GdNPF cDNA. The deduced human GdNPF amino cid sequence indicates that the protein is a member of a family of cell-derived protease inhibitors named rotease nexins. The development and the maintenance of the nervous system require a multiplicity of cellular interactions. In- creasing experimental evidence indicates that molecules derived from nonneuronal cells, especially glial cells, regulate survival and/or neurite outgrowth of neuronal cells (Monard et al., 1973; Barde et al., 1978; Lindsay, 1979; Sensenbrenner et al., 1980; Tanaka and Obata, 1982; Unsicker et al., 1984). A detailed characterization of the molecules involved is a prerequisite to understanding ?he nature of such cell-cell interactions. A 43 kd glia-derived neurite promoting factor (GdNPF) has been purified from medium conditioned by C6 rat glioma cells (Guenther et al., 1985). GdNPF is a potent serine protease inhibitor that forms an SDS-resistant com- plex with proteases such as urokinase, tissue plasmino- gen activator, thrombin, and trypsin (Monard et al., 1983; Guenther et al., 1985; Stone et al., submitted). Two other serine protease inhibitors (hirudin and the synthetic tri- peptide D-Phe-Pro-ArgCHI Monard, 1985). Purified GdNPF