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Research Projects

Kinetic Studies of Nuclear Import Pathways In Vivo

A quantitative single-cell measurement of nuclear import rates in vivo.
A quantitative single-cell measurement of nuclear import rates in vivo.

We are studying the kinetics and interactions between transport factors, transport cargoes, and NPC. Multiple karyopherins carry cargoes across the NPC; we have therefore developed an improved in vivo nuclear import assay to further characterize the reason why alternative pathways across the NPC should exist. We had previously described Kap123p as an alternative karyopherin that is involved in the nuclear import of ribosomal proteins.
Rout et al., 1997 (PDF)

Our work shows that cargoes of such small size require particularly efficient import, and that Kap123p provides this efficiency solely by virtue of its high abundance compared with other Kaps. Transport through the NPC proved to be saturable, and our import data fitted well to a pump-leak model of the import process.

Our eventual aim is to integrate our ultrastructural and biochemical studies to understand the molecular basis of the translocation of different transport factors across the NPC. These studies should enable us to reconstitute key reactions of these processes in vitro, study the high-resolution structures mediating the transport processes and test possible mechanistic models in vivo to understand the complete sequence of events during a transport cycle.