title: The RNA helicase RHAU (DHX36) suppresses expression of the transcription factor PITX1.
abstract: RNA Helicase associated with AU-rich element (RHAU) (DHX36) is a DEAH (Aspartic acid, Glumatic Acid, Alanine, Histidine)-box RNA helicase that can bind and unwind G4-quadruplexes in DNA and RNA. To detect novel RNA targets of RHAU, we performed an RNA co-immunoprecipitation screen and identified the PITX1 messenger RNA (mRNA) as specifically and highly enriched. PITX1 is a homeobox transcription factor with roles in both development and cancer. Primary sequence analysis identified three probable quadruplexes within the 3'-untranslated region of the PITX1 mRNA. Each of these sequences, when isolated, forms stable quadruplex structures that interact with RHAU. We provide evidence that these quadruplexes exist in the endogenous mRNA; however, we discovered that RHAU is tethered to the mRNA via an alternative non-quadruplex-forming region. RHAU knockdown by small interfering RNA results in significant increases in PITX1 protein levels with only marginal changes in mRNA, suggesting a role for RHAU in translational regulation. Involvement of components of the microRNA machinery is supported by similar and non-additive increases in PITX1 protein expression on Dicer and combined RHAU/Dicer knockdown. We also demonstrate a requirement of argonaute-2, a key RNA-induced silencing complex component, to mediate RHAU-dependent changes in PITX1 protein levels. These results demonstrate a novel role for RHAU in microRNA-mediated translational regulation at a quadruplex-containing 3'-untranslated region.
authors.affiliation: Department of Chemistry, University of Manitoba, Winnipeg, Manitoba, Canada, University of Manitoba, Winnipeg, Manitoba, Canada, Department of Biochemistry and Molecular Biology, University of British Columbia, V6T 1Z4 Vancouver, British Columbia, Canada and Department of Biochemistry and Medical Genetics, University of Manitoba, Winnipeg, Manitoba, Canada, R3T 2N2.
authors.forename: Evan P
authors.initials: EP
authors.lastname: Booy
authors.forename: Ryan
authors.initials: R
authors.lastname: Howard
authors.forename: Oksana
authors.initials: O
authors.lastname: Marushchak
authors.forename: Emmanuel O
authors.initials: EO
authors.lastname: Ariyo
authors.forename: Markus
authors.initials: M
authors.lastname: Meier
authors.forename: Stefanie K
authors.initials: SK
authors.lastname: Novakowski
authors.forename: Soumya R
authors.initials: SR
authors.lastname: Deo
authors.forename: Edis
authors.initials: E
authors.lastname: Dzananovic
authors.forename: Jörg
authors.initials: J
authors.lastname: Stetefeld
authors.forename: Sean A
authors.initials: SA
authors.lastname: McKenna
mesh_terms: D020413:3' Untranslated Regions; D060565:Argonaute Proteins / Q000378:metabolism; D001665:Binding Sites; D053487:DEAD-box RNA Helicases / Q000378:metabolism*; D054856:G-Quadruplexes; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D051761:Paired Box Transcription Factors / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D043244:Ribonuclease III / Q000037:antagonists & inhibitors
chemical_list: D020413:3' Untranslated Regions; C466422:AGO2 protein, human; D060565:Argonaute Proteins; D051761:Paired Box Transcription Factors; D012333:RNA, Messenger; C101485:homeobox protein PITX1; D043244:Ribonuclease III; C466988:DHX36 protein, human; D053487:DEAD-box RNA Helicases