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 whi ch plays an essential role in eukaryotic gene expression. As a karyoph ilic molecule, this cytosolic protein reaches its DNA-binding site thr ough 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 nu clear translocation of TBP, we detected ATP-dependent multimerization of TBP with atomic force microscopy. In physiological solutions contai ning ATP, 14-molecule multimers dissociated into four-molecule multime rs with a half-maximum dissociation constant of 10 mu M. Electrophysio logical experiments using isolated cell nuclei of cultured kidney cell s revealed that TBP translocates into the cell nucleus only in the pre sence of ATP. When ATP was replaced with its slowly hydrolysing analog ue, ATP[gamma-S] [i.e. adenosine 5'-o-(3-thiotriphosphate)], the aggre gates remained intact and nuclear translocation was not possible. Take n together, our investigations suggest that TBP exhibits ATPase activi ty similar to that observed in relation to molecular chaperons. This a ctivity secures physiological translocation of the transcription facto r into the nucleus.