The intranuclear mobility of messenger RNA binding proteins is ATP dependent and temperature sensitive.
After being released from transcription sites, messenger ribonucleoprotein particles (mRNPs) must reach the nuclear pore complexes in order to be translocated to the cytoplasm. Whether the intranuclear movement of mRNPs results largely from Brownian motion or involves molecular motors remains unknown. Here we have used quantitative photobleaching techniques to monitor the intranuclear mobility of protein components of mRNPs tagged with GFP. The results show that the diffusion coefficients of the poly(A)-binding protein II (PABP2) and the export factor TAP are significantly reduced when these proteins are bound to mRNP complexes, as compared with nonbound proteins. The data further show that the mobility of wild-type PABP2 and TAP, but not of a point mutant variant of PABP2 that fails to bind to RNA, is significantly reduced when cells are ATP depleted or incubated at 22 degrees C. Energy depletion has only minor effects on the intranuclear mobility of a 2,000-kD dextran (which corresponds approximately in size to 40S mRNP particles), suggesting that the reduced mobility of PABP2 and TAP is not caused by a general alteration of the nuclear environment. Taken together, the data suggest that the mobility of mRNPs in the living cell nucleus involves a combination of passive diffusion and ATP-dependent processes.
Calapez
Alexandre
A
Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, 1649-028 Lisboa, Portugal.
Pereira
Henrique M
HM
Calado
Angelo
A
Braga
José
J
Rino
José
J
Carvalho
Célia
C
Tavanez
João Paulo
JP
Wahle
Elmar
E
Rosa
Agostinho C
AC
Carmo-Fonseca
Maria
M
D021581:Active Transport, Cell Nucleus; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000502:physiology*; D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive; D002467:Cell Nucleus / Q000378:metabolism*; D003609:Dactinomycin / Q000494:pharmacology; D003911:Dextrans / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008164:Luminescent Proteins / Q000378:metabolism; D038761:Photobleaching; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D012680:Sensitivity and Specificity; D013696:Temperature; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D003911:Dextrans; D008164:Luminescent Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D049452:Green Fluorescent Proteins; D003609:Dactinomycin; D000255:Adenosine Triphosphate