title: Disrupting protein expression with Peptide Nucleic Acids reduces infection by obligate intracellular Rickettsia.
abstract: Peptide Nucleic Acids (PNAs) are single-stranded synthetic nucleic acids with a pseudopeptide backbone in lieu of the phosphodiester linked sugar and phosphate found in traditional oligos. PNA designed complementary to the bacterial Shine-Dalgarno or start codon regions of mRNA disrupts translation resulting in the transient reduction in protein expression. This study examines the use of PNA technology to interrupt protein expression in obligate intracellular Rickettsia sp. Their historically intractable genetic system limits characterization of protein function. We designed PNA targeting mRNA for rOmpB from Rickettsia typhi and rickA from Rickettsia montanensis, ubiquitous factors important for infection. Using an in vitro translation system and competitive binding assays, we determined that our PNAs bind target regions. Electroporation of R. typhi and R. montanensis with PNA specific to rOmpB and rickA, respectively, reduced the bacteria's ability to infect host cells. These studies open the possibility of using PNA to suppress protein synthesis in obligate intracellular bacteria.
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Pelc
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: McClure
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Simran J
authors.initials: SJ
authors.lastname: Kaur
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Khandra T
authors.initials: KT
authors.lastname: Sears
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: M Sayeedur
authors.initials: MS
authors.lastname: Rahman
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Shane M
authors.initials: SM
authors.lastname: Ceraul
mesh_terms: D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015153:Blotting, Western; D057846:Cell-Penetrating Peptides / Q000494:pharmacology; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000382:microbiology; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D051379:Mice; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012281:Rickettsia / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D012282:Rickettsia Infections / Q000188:drug therapy / Q000235:genetics / Q000382:microbiology; D014709:Vero Cells
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D057846:Cell-Penetrating Peptides; D020135:Peptide Nucleic Acids; D012333:RNA, Messenger; C491314:RickA protein, Rickettsia; C080205:rOmpA, Rickettsia