Dual roles of RNA helicase A in CREB-dependent transcription.
RNA helicase A (RHA) is a member of an ATPase/DNA and RNA helicase family and is a homologue of Drosophila maleless protein (MLE), which regulates X-linked gene expression. RHA is also a component of holo-RNA polymerase II (Pol II) complexes and recruits Pol II to the CREB binding protein (CBP). The ATPase and/or helicase activity of RHA is required for CREB-dependent transcription. To further understand the role of RHA on gene expression, we have identified a 50-amino-acid transactivation domain that interacts with Pol II and termed it the minimal transactivation domain (MTAD). The protein sequence of this region contains six hydrophobic residues and is unique to RHA homologues and well conserved. A mutant with this region deleted from full-length RHA decreased transcriptional activity in CREB-dependent transcription. In addition, mutational analyses revealed that several tryptophan residues in MTAD are important for the interaction with Pol II and transactivation. These mutants had ATP binding and ATPase activities comparable to those of wild-type RHA. A mutant lacking ATP binding activity was still able to interact with Pol II. In CREB-dependent transcription, the transcriptional activity of each of these mutants was less than that of wild-type RHA. The activity of the double mutant lacking both functions was significantly lower than that of each mutant alone, and the double mutant had a dominant negative effect. These results suggest that RHA could independently regulate CREB-dependent transcription either through recruitment of Pol II or by ATP-dependent mechanisms.
Aratani
S
S
Institute of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan.
Fujii
R
R
Oishi
T
T
Fujita
H
H
Amano
T
T
Ohshima
T
T
Hagiwara
M
M
Fukamizu
A
A
Nakajima
T
T
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001324:Autoantigens / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D001665:Binding Sites; D017173:Caenorhabditis elegans; D017124:Conserved Sequence; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D053487:DEAD-box RNA Helicases; D006801:Humans; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D012319:RNA Polymerase II / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D001324:Autoantigens; D017362:Cyclic AMP Response Element-Binding Protein; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate; D012319:RNA Polymerase II; D000251:Adenosine Triphosphatases; C098487:DHX9 protein, human; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases