title: Pseudouridine andN6-methyladenosine modifications weaken PUF protein/RNA interactions.
abstract: RNA modifications are ubiquitous in biology, with over 100 distinct modifications. While the vast majority were identified and characterized on abundant noncoding RNA such as tRNA and rRNA, the advent of sensitive sequencing-based approaches has led to the discovery of extensive and regulated modification of eukaryotic messenger RNAs as well. The two most abundant mRNA modifications-pseudouridine (Ψ) andN6-methyladenosine (m6A)-affect diverse cellular processes including mRNA splicing, localization, translation, and decay and modulate RNA structure. Here, we test the hypothesis that RNA modifications directly affect interactions between RNA-binding proteins and target RNA. We show that Ψ and m6A weaken the binding of the human single-stranded RNA binding protein Pumilio 2 (hPUM2) to its consensus motif, with individual modifications having effects up to approximately threefold and multiple modifications giving larger effects. While there are likely to be some cases where RNA modifications essentially fully ablate protein binding, here we see modest responses that may be more common. Such modest effects could nevertheless profoundly alter the complex landscape of RNA:protein interactions, and the quantitative rather than qualitative nature of these effects underscores the need for quantitative, systems-level accounting of RNA:protein interactions to understand post-transcriptional regulation.
authors.affiliation: Department of Biochemistry, Stanford University, Stanford, California 94305, USA.
authors.forename: Pavanapuresan P
authors.identifier: 0000-0001-6764-3666
authors.initials: PP
authors.lastname: Vaidyanathan
authors.affiliation: Department of Biochemistry, Stanford University, Stanford, California 94305, USA.
authors.forename: Ishraq
authors.initials: I
authors.lastname: AlSadhan
authors.affiliation: Department of Chemistry, Duke University, Durham, North Carolina 27708, USA.
authors.forename: Dawn K
authors.initials: DK
authors.lastname: Merriman
authors.affiliation: Department of Chemistry, Duke University, Durham, North Carolina 27708, USA.|Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Hashim M
authors.initials: HM
authors.lastname: Al-Hashimi
authors.affiliation: Department of Biochemistry, Stanford University, Stanford, California 94305, USA.|Departments of Chemical Engineering and Chemistry, Stanford University, Stanford, California 94305, USA.|Stanford ChEM-H (Chemistry, Engineering, and Medicine for Human Health), Stanford University, Stanford, California 94305, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Herschlag
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D011485:Protein Binding; D011560:Pseudouridine / Q000378:metabolism*; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*
chemical_list: C471243:PUM2 protein, human; D016601:RNA-Binding Proteins; D011560:Pseudouridine; D012313:RNA; C010223:N-methyladenosine; D000241:Adenosine
keywords: N6-methyladenosine; PUMILIO; RNA binding proteins; RNA–protein interactions; epitranscriptomics; pseudouridine