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Top3β is an RNA topoisomerase that works with fragile X syndrome protein to promote synapse formation

Abstract

Topoisomerases are crucial for solving DNA topological problems, but they have not been linked to RNA metabolism. Here we show that human topoisomerase 3β (Top3β) is an RNA topoisomerase that biochemically and genetically interacts with FMRP, a protein that is deficient in fragile X syndrome and is known to regulate the translation of mRNAs that are important for neuronal function, abnormalities of which are linked to autism. Notably, the FMRP-Top3β interaction is abolished by a disease-associated mutation of FMRP, suggesting that Top3β may contribute to the pathogenesis of mental disorders. Top3β binds multiple mRNAs encoded by genes with neuronal functions linked to schizophrenia and autism. Expression of one such gene, that encoding protein tyrosine kinase 2 (ptk2, also known as focal adhesion kinase or FAK), is reduced in the neuromuscular junctions of Top3β mutant flies. Synapse formation is defective in Top3β mutant flies and mice, as well as in FMRP mutant flies and mice. Our findings suggest that Top3β acts as an RNA topoisomerase and works with FMRP to promote the expression of mRNAs that are crucial for neurodevelopment and mental health.

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Figure 1: Top3β and TDRD3 form a complex that associates with FMRP; this association is disrupted by a patient-derived point mutation or substitution of methylated arginine residues in FMRP.
Figure 2: TDRD3 acts as a bridge connecting Top3β and FMRP.
Figure 3: Top3β associates with TDRD3 and FMRP in stress granules and polyribosomes.
Figure 4: Top3β has RNA topoisomerase activity that depends on a conserved RGG RNA-binding motif.
Figure 5: Top3β binds coding regions of mRNAs, and its bound mRNAs are enriched with matching FMRP targets.
Figure 6: Top3β and Tdrd3 mutations modify Fmr1 function in Drosophila eyes.
Figure 7: Drosophila Top3β and Fmr1 work in a common pathway to promote the formation of NMJs.
Figure 8: Drosophila Top3β and Fmr1 work in the same pathway to promote the expression of ptk2.

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Acknowledgements

We thank T. Hsieh (Duke University) for Drosophila Top3β reagents; J.C. Wang (Harvard University) for Top3β knockout mice; D. Zarnescu (University of Illinois), T. Jongens (University of Pennsylvania) and G. Dreyfuss (University of Pennsylvania) for dfmr1 fly strains and antibodies; S. Warren (Emory University), S. Ceman (University of Illinois) and Y. Feng (Emory University) for vectors of FMR1 variants; U. Fischer (University of Wuerzburg) for vectors of TDRD3 and FMR1 orthologs; A. Hoogeveen (Erasmus University) for FXR1 and FXR2 antibodies; R. Hynes (Massachusetts Institute of Technology) and R. Palmer (Umea University) for FAK reagents; T. Enomoto (Tohoku University) for Top3b−/− DT40 cells; and R. Hanai (Rikkyo University) for the Top3β vector. We thank E. Chen, D.J. Pan and Y. Feng for advice and assistance, S.K. Lee for assistance and D. Schlessinger for support and critical reading of the manuscript. This work is supported in part by the Intramural Research Program of the NIA (Z01 AG000657-08), the NIH, the Johns Hopkins Center for Neuroscience Research (NS050274), Canadian Institutes of Health research grant MOP-79368 (to G.W.B.), the National Basic Research Program of China (2013CB911002) and the National Natural Science Foundation of China (31271435). This study used the high-performance computational capabilities of the Biowulf Linux cluster at the NIH, Bethesda, Maryland, USA (http://biowulf.nih.gov).

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D.X., W.S., R.G., Y.X., W.P., J.S., Jay Yang, S.S., Jiandong Yang, D.F., J.L.M., Y.P., J.M., S.R.J. and S.M. conducted experiments. A.C.S., T.E.L., G.W.B., M.S.H.K., M.G., S.Z. and W.W. supervised the project. A.S., Y.Q., D.X., W.S., R.G., Y.X., W.P., J.S., Jay Yang, S.S., A.C.S., T.E.L., G.W.B., M.S.H.K., M.G., S.Z. and W.W. analyzed the data. D.X., W.S., M.G., S.Z. and W.W. wrote the manuscript with input from the other authors.

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Correspondence to Sige Zou or Weidong Wang.

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Xu, D., Shen, W., Guo, R. et al. Top3β is an RNA topoisomerase that works with fragile X syndrome protein to promote synapse formation. Nat Neurosci 16, 1238–1247 (2013). https://doi.org/10.1038/nn.3479

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