title: Biophysical Characterization of G-Quadruplex Recognition in the PITX1 mRNA by the Specificity Domain of the Helicase RHAU.
abstract: Nucleic acids rich in guanine are able to fold into unique structures known as G-quadruplexes. G-quadruplexes consist of four tracts of guanylates arranged in parallel or antiparallel strands that are aligned in stacked G-quartet planes. The structure is further stabilized by Hoogsteen hydrogen bonds and monovalent cations centered between the planes. RHAU (RNA helicase associated with AU-rich element) is a member of the ATP-dependent DExH/D family of RNA helicases and can bind and resolve G-quadruplexes. RHAU contains a core helicase domain with an N-terminal extension that enables recognition and full binding affinity to RNA and DNA G-quadruplexes. PITX1, a member of the bicoid class of homeobox proteins, is a transcriptional activator active during development of vertebrates, chiefly in the anterior pituitary gland and several other organs. We have previously demonstrated that RHAU regulates PITX1 levels through interaction with G-quadruplexes at the 3'-end of the PITX1 mRNA. To understand the structural basis of G-quadruplex recognition by RHAU, we characterize a purified minimal PITX1 G-quadruplex using a variety of biophysical techniques including electrophoretic mobility shift assays, UV-VIS spectroscopy, circular dichroism, dynamic light scattering, small angle X-ray scattering and nuclear magnetic resonance spectroscopy. Our biophysical analysis provides evidence that the RNA G-quadruplex, but not its DNA counterpart, can adopt a parallel orientation, and that only the RNA can interact with N-terminal domain of RHAU via the tetrad face of the G-quadruplex. This work extends our insight into how the N-terminal region of RHAU recognizes parallel G-quadruplexes.
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Emmanuel O
authors.initials: EO
authors.lastname: Ariyo
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Evan P
authors.initials: EP
authors.lastname: Booy
authors.affiliation: School of Biosciences, University of Birmingham, Birmingham B152TT, United Kingdom.
authors.forename: Trushar R
authors.initials: TR
authors.lastname: Patel
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Edis
authors.initials: E
authors.lastname: Dzananovic
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Ewan K
authors.initials: EK
authors.lastname: McRae
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Markus
authors.initials: M
authors.lastname: Meier
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.|Manitoba Institute for Materials, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Kevin
authors.initials: K
authors.lastname: McEleney
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.|Manitoba Institute for Materials, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.|Department of Biochemistry and Medical Genetics, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Jorg
authors.initials: J
authors.lastname: Stetefeld
authors.affiliation: Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.|Manitoba Institute for Materials, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.|Department of Biochemistry and Medical Genetics, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada.
authors.forename: Sean A
authors.initials: SA
authors.lastname: McKenna
mesh_terms: D063307:AU Rich Elements; D001703:Biophysics; D002942:Circular Dichroism; D053487:DEAD-box RNA Helicases / Q000502:physiology*; D004247:DNA / Q000378:metabolism; D054856:G-Quadruplexes; D006801:Humans; D051761:Paired Box Transcription Factors / Q000378:metabolism; D011485:Protein Binding; D012333:RNA, Messenger / Q000032:analysis*
chemical_list: D051761:Paired Box Transcription Factors; D012333:RNA, Messenger; C101485:homeobox protein PITX1; D004247:DNA; C466988:DHX36 protein, human; D053487:DEAD-box RNA Helicases