title: Functional amyloids as inhibitors of plasmid DNA replication.
abstract: DNA replication is tightly regulated to constrain the genetic material within strict spatiotemporal boundaries and copy numbers. Bacterial plasmids are autonomously replicating DNA molecules of much clinical, environmental and biotechnological interest. A mechanism used by plasmids to prevent over-replication is 'handcuffing', i.e. inactivating the replication origins in two DNA molecules by holding them together through a bridge built by a plasmid-encoded initiator protein (Rep). Besides being involved in handcuffing, the WH1 domain in the RepA protein assembles as amyloid fibres upon binding to DNA in vitro. The amyloid state in proteins is linked to specific human diseases, but determines selectable and epigenetically transmissible phenotypes in microorganisms. Here we have explored the connection between handcuffing and amyloidogenesis of full-length RepA. Using a monoclonal antibody specific for an amyloidogenic conformation of RepA-WH1, we have found that the handcuffed RepA assemblies, either reconstructed in vitro or in plasmids clustering at the bacterial nucleoid, are amyloidogenic. The replication-inhibitory RepA handcuff assembly is, to our knowledge, the first protein amyloid directly dealing with DNA. Built on an amyloid scaffold, bacterial plasmid handcuffs can bring a novel molecular solution to the universal problem of keeping control on DNA replication initiation.
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Molina-García
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: Fátima
authors.initials: F
authors.lastname: Gasset-Rosa
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Moreno-Del Álamo
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: M Elena
authors.initials: ME
authors.lastname: Fernández-Tresguerres
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: Susana
authors.initials: S
authors.lastname: Moreno-Díaz de la Espina
authors.affiliation: Max Planck Institute for Molecular Genetics, D14195 Berlin, Germany.
authors.forename: Rudi
authors.initials: R
authors.lastname: Lurz
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas - CSIC, E28040 Madrid, Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Giraldo
mesh_terms: D000682:Amyloid / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D000906:Antibodies / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D004261:DNA Replication / Q000187:drug effects*; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008854:Microscopy, Electron; D010957:Plasmids / Q000187:drug effects / Q000235:genetics*; D011487:Protein Conformation; D018741:Replication Origin; D015534:Trans-Activators / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*
chemical_list: D000682:Amyloid; D000906:Antibodies; D004269:DNA, Bacterial; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases