The Karyopherinβ family of proteins (Kapβs; Importins/Exportins) mediate the majority of macromolecular traffic between the nucleus and the cytoplasm. Kapβs bind specific sets of substrates through nuclear localization or nuclear export signals (NLSs or NESs), and transport them through the nuclear pore complex. Research in the Chook lab is directed towards understanding the physical and cellular mechanisms of Kapβs. Our long-term goals are to understand the macromolecular nuclear traffic patterns coordinated by the 19 human Kapβs and learn how these patterns contribute to overall cellular organization. It is unclear why the cell needs so many different nuclear importers and exporters. Are their collections of substrates random except for a common signal? Or do individual Kapβs transport substrates with distinct cellular functions? What is the extent of substrate overlap between pathways? Our research aims to answer these fundamental questions.
To achieve our goals, we have initially focused on Kapβ2/Transportin as a model system. Most importantly, using structural (primarily X-ray crystallography), biochemical and bioinformatics approaches, we have defined a set of rules for NLS recognition by Kapβ2 and used these to discover many new candidate substrates. This work revealed the first new class of NLS (termed PY-NLS) in 25 years. It also established a general framework for future discovery of nucleocytoplasmic targeting signals. More generally, we have discovered complex signals using a collection of physical rules rather than specific sequence motifs alone, and this concept could be expanded across organelle systems to study the numerous obscure targeting signals in eukaryotic cells. Subsequent comparative structural analysis of Kapβ2 bound to diverse PY-NLSs confirmed our NLS rules. Biophysical studies of these complexes also enabled us to design a nuclear import inhibitor that specifically inhibits Kapβ2-mediated nuclear import in cells. More recently, extensive thermodynamic analyses have revealed a set of biophysical properties that govern the affinity of PY-NLS. Finally, we have determined the mechanism by which RanGTP releases substrates from Kapβ2.
Our lab is taking a broadly multidisciplinary approach to study substrate interactions in all Kapβ pathways. We are using Kapβ2 as a model system to develop approaches to map substrate repertoire, and discover and understand functional program(s) for a Kapβ pathway. We have already established strategies to develop predictive rules for substrates, but suspect that the 81 Kapβ2 substrates we have predicted constitute only a small fraction of all Kapβ2 substrates, and a more complete mapping is necessary for systems-level analysis. We will deepen our biophysical understanding of NLS recognition and in vivo nuclear import to develop a computational method to comprehensively identify Kapβ2 substrates, and to analyze the trafficking networks for functional linkages. Ultimately, we will study how nuclear trafficking impacts on the suggested biological processes. Our Kapβ2 work will establish a framework for discovery of nuclear transport signals, generation of comprehensive traffic maps and protein networks analysis that we will apply to all Kapβ pathways.
RESEARCH INTERESTS
Protein transport, nucleocytoplasmic transport
Structural Biology
Biophysics
Bioinformatics
Cell Biology
SIGNIFICANT PUBLICATIONS
Cansizoglu, A. E. and Chook, Y. M., "Conformational heterogeneity of Karyopherin-beta2 is segmental." Structure, 15(11):1431-1441, November 2007
Cansizoglu, A. E., Lee, B. J., Zhang, Z. C., Fontoura, B. M. A. and Chook, Y. M., "Structure-based design of a pathway-specific nuclear import inhibitor." Nat. Struc. & Mol. Biol., 14(5):452-454, May 2007
Lee, B. J., Cansizoglu, A. E., Suel, K. E., Louis, T. H., Zichao Zhang and Chook, Y. M., "Rules for nuclear localization sequence recognition by Karyopherinβ2." Cell, 126:543-558, August 2006
Chook, Y.M., Jung, A., Rosen, M.K. and Blobel, G., "Uncoupling Ran binding and substrate dissociation in the Karyopherinβ2 nuclear import pathway." Biochemistry, 41:6955-6966, June 2002
Chook, Y.M. and Blobel, G., "Structure of the nuclear transport complex karyopherinβ2-Ran.GppNHp." Nature, 399:230-237, May 1999
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