Atomic force microscopy visualizes ATP-dependent dissociation of multimeric TATA-binding protein before translocation into the cell nucleus.
The TATA-binding protein (TBP) is a universal transcription factor which plays an essential role in eukaryotic gene expression. As a karyophilic molecule, this cytosolic protein reaches its DNA-binding site through the transport channel of the nuclear pore complex. As occurs with other major cellular proteins, TBP forms multimers in solution, which is a limiting factor for nuclear translocation. While studying the nuclear translocation of TBP, we detected ATP-dependent multimerization of TBP with atomic force microscopy. In physiological solutions containing ATP, 14-molecule multimers dissociated into four-molecule multimers with a half-maximum dissociation constant of 10 microM. Electrophysiological experiments using isolated cell nuclei of cultured kidney cells revealed that TBP translocates into the cell nucleus only in the presence of ATP. When ATP was replaced with its slowly hydrolysing analogue, ATP[gamma-S] [i.e. adenosine 5'-o-(3-thiotriphosphate)], the aggregates remained intact and nuclear translocation was not possible. Taken together, our investigations suggest that TBP exhibits ATPase activity similar to that observed in relation to molecular chaperons. This activity secures physiological translocation of the transcription factor into the nucleus.
Oberleithner
H
H
Department of Physiology, University of Würzburg School of Medicine, Röntgenring 9, D-97070 Würzburg, Germany.
Schneider
S
S
Bustamante
J O
JO
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D008564:Membrane Potentials / Q000187:drug effects; D018625:Microscopy, Atomic Force; D009685:Nuclear Envelope / Q000187:drug effects; D011487:Protein Conformation; D012996:Solutions; D035181:TATA-Box Binding Protein; D014157:Transcription Factors / Q000378:metabolism*
D004268:DNA-Binding Proteins; D012996:Solutions; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate