Identification of functional domains involved in BTG1 cell localization

We have previously shown that BTG1 stimulates myoblast differentiation, In addition, this protein displays a major nuclear localization in confluent m yoblasts, decreasing during the early steps of differentiation, and is esse ntially detected in the cytoplasm of mature myotubes. To identify the domai ns involved in the cellular trafficking of BTG1, we observed the localizati on of several BTG1 sequences fused to beta Galactosidase. The highly conser ved B box among all members of the BTG family induces a significant nuclear localization of the beta Gal moiety, enhanced by presence of the BTG1 carb oxy-terminal sequence. In addition, a functional Nuclear Export Signal (NES ) overlaps the B box. Moreover, presence of the first 43 NH2-terminal amino acids reduced the nuclear localization of each chimeric protein tested. La st, the BTG1 amino-terminal domain hears an LxxLL, motif favouring nuclear accumulation, and another region encompassing the A box inhibiting nuclear localization. In contrast to a BTG1 mutant exclusively localized in the cyt oplasm, transient expression of a mutant displaying a nuclear Localization enhanced myoblasts withdrawal from the cell cycle and terminal differentiat ion, thus mimicking the myogenic influence of BTG1, In conclusion, several regions of BTG1 are implicated in its cellular localization, and BTG1 myoge nic activity is induced at the nuclear level.