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11:32
Analysis of mRNA Nuclear Export Kinetics in Mammalian Cells by Microinjection
Published on: December 4, 2010
A supraphysiological nuclear export signal is required for parvovirus nuclear export
Dieuwke Engelsma1, Noelia Valle, Alexander Fish
1Department of Tumor Biology, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands.
Molecular Biology of the Cell
|April 4, 2008
Summary
Supraphysiological nuclear export signals (NESs) exist in nature. The Minute Virus of Mice NS2 protein uses a supraphysiological NES for efficient viral nuclear export and replication.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Nuclear export signal (NES) proteins are exported from the nucleus via CRM1.
- Optimal function of NESs requires low affinity for CRM1.
- Artificial supraphysiological NESs with higher CRM1 affinity were previously generated.
Purpose of the Study:
- To identify a naturally occurring supraphysiological NES in an endogenous protein.
- To investigate the role of this NES in viral replication and nuclear export.
Main Methods:
- Identification of a supraphysiological NES in the NS2 protein of Minute Virus of Mice (MVM).
- Analysis of NS2-CRM1 interaction with and without RanGTP.
- Mutation of a key residue in the NS2 NES to assess its effect on CRM1 affinity and viral function.
- Observation of colocalization of NS2, CRM1, and MVM virions in infected cells.
Main Results:
- A supraphysiological NES was identified in the MVM NS2 protein.
- NS2 interacts with CRM1, with RanGTP enhancing complex stability.
- Mutation of a hydrophobic residue reduced NS2 NES affinity to regular levels, impairing viral nuclear export and productivity.
- NS2, CRM1, and MVM virions colocalized in infected cells, dependent on the supraphysiological NS2 NES.
Conclusions:
- Supraphysiological NESs are naturally occurring.
- The supraphysiological NS2 NES is critical for active nuclear export of MVM particles prior to cell lysis.
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