title: Recognition of Specified RNA Modifications by the Innate Immune System.
abstract: Microbial nucleic acids have been described as important activators of human innate immune responses by triggering so-called pattern recognition receptors (PRRs) that are expressed on innate immune cells, including plasmacytoid dendritic cells and monocytes. Although host and microbial nucleic acids share pronounced chemical and structural similarities, they significantly differ in their posttranscriptional modification profile, allowing the host to discriminate between self and nonself. In this regard, ribose 2'-O-methylation has been discovered as suppressor of RNA-induced PRR activation. Although 2'-O-methylation occurs with higher frequencies in eukaryotic than in prokaryotic RNA, the immunosuppressive properties of 2'-O-methylated nucleotides may be misused by certain bacteria as immune evasion mechanism. In the course of identifying inhibitory RNA modifications, our groups have synthesized and comparatively analyzed a series of differentially modified RNAs, so-called modivariants, for their immune stimulatory capacities. In this chapter, we will detail the protocols for the design and synthesis of RNA modivariants by molecular cut-and-paste techniques (referred to as molecular surgery) and describe testing of their immune stimulatory properties upon transfection into peripheral blood mononuclear cells.
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg, Heidelberg, Germany.
authors.forename: Tatjana
authors.initials: T
authors.lastname: Eigenbrod
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Mainz, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Keller
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Mainz, Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Kaiser
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg, Heidelberg, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Rimbach
authors.affiliation: Department of Infectious Diseases, Medical Microbiology and Hygiene, University of Heidelberg, Heidelberg, Germany.
authors.forename: Alexander H
authors.initials: AH
authors.lastname: Dalpke
authors.affiliation: Institute of Pharmacy and Biochemistry, Johannes Gutenberg-University of Mainz, Mainz, Germany. Electronic address: mhelm@uni-mainz.de.
authors.forename: Mark
authors.initials: M
authors.lastname: Helm
mesh_terms: D003713:Dendritic Cells / Q000276:immunology; D006801:Humans; D007113:Immunity, Innate; D007963:Leukocytes, Mononuclear / Q000737:chemistry / Q000276:immunology; D008745:Methylation; D009000:Monocytes / Q000276:immunology; D009696:Nucleic Acids / Q000276:immunology*; D012313:RNA / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics*; D012323:RNA Processing, Post-Transcriptional / Q000276:immunology*; D012329:RNA, Bacterial / Q000737:chemistry / Q000276:immunology; D051192:Receptors, Pattern Recognition / Q000096:biosynthesis* / Q000276:immunology; D012266:Ribose / Q000737:chemistry / Q000276:immunology
chemical_list: D009696:Nucleic Acids; D012329:RNA, Bacterial; D051192:Receptors, Pattern Recognition; D012313:RNA; D012266:Ribose
keywords: Catalytic DNA; DNA splint; DNAzyme; Immune stimulation; Ligation; Modivariant; Molecular surgery; Plasmacytoid dendritic cells; RNA cleavage; tRNA