C. elegans CEBP-1 is a C/EBP bZIP transcription factor and essential for axon regeneration. Previous studies have focused on the role of bZIP domain at the C-terminus of CEBP-1 in axon regeneration (Yan et al., 2009 Cell). However, CEBP-1 also has a long N-terminal fragment that contains no known protein motifs and how it is involved in CEBP-1 function remains unknown. Here, we report novel structural motifs in the N-terminus of CEBP-1 for axon regeneration and nuclear import. Using two types of forward genetic screens, we have identified a unique domain in the N-terminus that is critical for CEBP-1's in vivo functions. This N' domain also has a predicted propensity to form alpha helices. Furthermore, using yeast two-hybrid screening with N-terminal CEBP-1 fragments as baits, we have identified IMA-3/Importin-? as a CEBP-1 binding partner. In transgenic expression studies, we characterized the subcellular localization of CEBP-1 fragments and identified three nuclear localization signals (NLS) in CEBP-1, one which interacts with IMA-3. Lastly, we showed that
ima-3 is required for axon regeneration. Taken together, our findings uncover new protein structural elements for CEBP-1, and further reinforce the importance of Importin-mediated nuclear import system for promoting axon regeneration. Yan, D., Wu, Z., Chisholm, A.D., and Jin, Y. (2009). The DLK-1 kinase promotes mRNA stability and local translation in C. elegans synapses and axon regeneration. Cell 138, 1005-1018.