Abstract:
:We have investigated classical nuclear localization sequence (NLS) mediated protein trafficking by measuring biomolecular dynamics within living cells using two-photon fluorescence correlation spectroscopy. By directly observing the behavior of specific molecules in their native cellular environment, it is possible to uncover functional details that are not apparent from traditional biochemical investigations or functional assays. We show that the intracellular mobility of NLS cargoes and their import receptor proteins, karyopherin-alpha and karyopherin-beta, can be robustly measured and that quantitative comparison of intracellular diffusion coefficients provides new insights into nuclear transport mechanisms. Import cargo complexes are assembled throughout the cytoplasm, and their diffusion is slower than predicted by molecular weight due to specific interactions. Analysis of NLS cargo diffusion in the cytoplasm indicates that these interactions are likely disrupted by NLS cargo binding. Our results suggest that delivery of import receptors and NLS cargoes to nuclear pores may complement selective translocation through the pores as a functional mechanism for regulating transport of proteins into the nucleus.
journal_name
Biophys Jjournal_title
Biophysical journalauthors
Wu J,Corbett AH,Berland KMdoi
10.1016/j.bpj.2009.01.050subject
Has Abstractpub_date
2009-05-06 00:00:00pages
3840-9issue
9eissn
0006-3495issn
1542-0086pii
S0006-3495(09)00583-9journal_volume
96pub_type
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