|Budget Amount *help
¥3,300,000 (Direct Cost : ¥3,300,000)
Fiscal Year 1998 : ¥1,200,000 (Direct Cost : ¥1,200,000)
Fiscal Year 1997 : ¥2,100,000 (Direct Cost : ¥2,100,000)
Nuclear import of proteins arc initiated by nuclear localization signal (NLS)-dependent complex formation, termed the nuclear pore-targeting complex (PTAC), in the cytoplasm. PTAC is a stable protein complex composed of a karyophilic protein and two cytosolic components termed PTAC58/importin α and PTAC97/importin β. Importin α directly binds to NLS of karyophile and importin β provides nuclear pore complex (NPC) binding site for PTAC. We analyzed the properties of importin α and importin β, and obtained the following results : 1. We have identified three additional mouse genes homologous to the known importin α, and showed that the mouse importin α can be classified into three subfamilies (Rch1, NPI1, and Qip1), showing approximately 50% amino acid identity to one another. 2. We showed that importin α family proteins show some specificities in the recognition of substrates. For example, Qip1 recognizes a very restricted NLS, requiring both the upstream sequence and basic core sequence
of Q1 ATPase for its substrate recognition. The other evidence is that tyrosine phosphorylated Stat his recognized by NPI1, but not Rch1. Stat 1 apparently does not possess typical basic-type NLS, and it binds to different portion of NPI1 from those of SV40 T-antigen basic NLS binding site. 3. We examined the behavior and functional domain of importin β Importin β rapidly shuttles between the cytoplasm and the nucleus through NPCs. The use of deletion mutants reveals that NPC-translocation of importin β requires neither Ran- nor importin α-binding but only the NPC-binding domain of this molecule. NPC translocation of importin β was found to require neither Ran nor its GTP hydrolysis. Furthermore, importin β possesses ability to translocate NPC on its own. This could be a common feature of transport factors that carries certain cargos through NPCs into and out of the nucleus. 4. We examined the nuclear import of β-catenin and found that b-catenin can translocate NPCs on it own without requiring soluble factors including Ran. Β-catenin is likely to share the same property with importin β for NPC translocation.