title: TMPyP4 porphyrin distorts RNA G-quadruplex structures of the disease-associated r(GGGGCC)n repeat of the C9orf72 gene and blocks interaction of RNA-binding proteins.
abstract: Certain DNA and RNA sequences can form G-quadruplexes, which can affect genetic instability, promoter activity, RNA splicing, RNA stability, and neurite mRNA localization. Amyotrophic lateral sclerosis and frontotemporal dementia can be caused by expansion of a (GGGGCC)n repeat in the C9orf72 gene. Mutant r(GGGGCC)n- and r(GGCCCC)n-containing transcripts aggregate in nuclear foci, possibly sequestering repeat-binding proteins such as ASF/SF2 and hnRNPA1, suggesting a toxic RNA pathogenesis, as occurs in myotonic dystrophy. Furthermore, the C9orf72 repeat RNA was recently demonstrated to undergo the noncanonical repeat-associated non-AUG translation (RAN translation) into pathologic dipeptide repeats in patient brains, a process that is thought to depend upon RNA structure. We previously demonstrated that the r(GGGGCC)n RNA forms repeat tract length-dependent G-quadruplex structures that bind the ASF/SF2 protein. Here we show that the cationic porphyrin (5,10,15,20-tetra(N-methyl-4-pyridyl) porphyrin (TMPyP4)), which can bind some G-quadruplex-forming sequences, can bind and distort the G-quadruplex formed by r(GGGGCC)8, and this ablates the interaction of either hnRNPA1 or ASF/SF2 with the repeat. These findings provide proof of concept that nucleic acid binding small molecules, such as TMPyP4, can distort the secondary structure of the C9orf72 repeat, which may beneficially disrupt protein interactions, which may ablate either protein sequestration and/or RAN translation into potentially toxic dipeptides. Disruption of secondary structure formation of the C9orf72 RNA repeats may be a viable therapeutic avenue, as well as a means to test the role of RNA structure upon RAN translation.
authors.affiliation: From the Graduate Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario M5S 3M2.
authors.forename: Bita
authors.initials: B
authors.lastname: Zamiri
authors.forename: Kaalak
authors.initials: K
authors.lastname: Reddy
authors.forename: Robert B
authors.initials: RB
authors.lastname: Macgregor
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Pearson
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics*; D001483:Base Sequence; D000073885:C9orf72 Protein; D042622:DNA Repeat Expansion / Q000235:genetics*; D057180:Frontotemporal Dementia / Q000235:genetics*; D054856:G-Quadruplexes / Q000187:drug effects*; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism; D011166:Porphyrins / Q000737:chemistry / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D011506:Proteins / Q000235:genetics*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D000068103:Serine-Arginine Splicing Factors
chemical_list: D000073885:C9orf72 Protein; C568651:C9orf72 protein, human; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D009687:Nuclear Proteins; D011166:Porphyrins; D011506:Proteins; D016601:RNA-Binding Proteins; C000619656:hnRNPA1 protein, human; D000068103:Serine-Arginine Splicing Factors; C021096:tetra(4-N-methylpyridyl)porphine
keywords: ASF/SF2; Amyotrophic Lateral Sclerosis (Lou Gehrig Disease); C9orf72; G-quadruplex; RNA; RNA Structure; RNA-Protein Interaction; RNA-binding Protein; TMPyP4; hnRNPA1