title: Inhibition of innate immune activation by modifications in bacterial RNA
creator: Rimbach, Katharina 
subject: ddc-570
subject: 570 Life sciences
description: Modification of RNA is an important function in the immunological discrimination between self and non-self RNA. Naturally occurring RNA modifications have been identified that suppress activation of the innate immune response, e.g. 2’O-methylation of the ribose backbone. It has already been shown that incorporation of 2’O-methylation in siRNA abrogates TLR7-dependent interferon-alpha production in human plasmacytoid dendritic cells (pDCs). In the present study, a native bacterial tRNA with 2’O-methylation at position G18 was analysed. It was found that the 2’O-methylation not only made the tRNA non-stimulatory, but also led to dominant inhibitory effects. Thus, this modification impaired immunostimulation by RNA species that would otherwise have been stimulatory. Further analysis revealed that not only was interferon alpha production by pDCs via TLR7 suppressed, but also secretion of pro-inflammatory cytokines by monocytes, including IL12p40 and TNF. Monocytes express a different subset of nucleic acid-sensing receptors, indicating that 2’O-methylated RNA influences the stimulation of different receptors. Analysis of other TLR ligands showed that the inhibitory effect was specific for stimulation by RNA but it was not observed for R848, a synthetic small molecule agonist of TLR7/8, or other TLR ligands. Investigation of the signalling pathway in response to RNA stimulation showed that modified, inhibitory RNA suppresses TLR stimulation at proximal levels. Preliminary binding studies indicated that 2’O-methylated RNA can bind directly to TLR7; it does not induce signalling but displaces unmethylated stimulatory RNA. As 2’O-methylation of tRNA at position G18 is found in tRNAs from many species and the neighbouring sequences are conserved, the influence of location and the whole sequence motif itself were investigated. The location of the   2’O-modification turned out to be of minor importance, but the inhibitory capacity of modified tRNA was influenced by the sequence of the motif. The sequence motif DmR was found to be necessary for immunosilencing of the tRNA, where D is a base other than cytosine with a 2’O-methyl position in the ribose of the nucleoside and R is a purine (adenine or guanine) immediately upstream of the methylation (position +1).  Together, the results show that 2’O-methylation of RNA in a defined sequence motif abrogates immunostimulation in different cell types. Naturally occurring RNA modifications serve to distinguish self RNA from foreign RNA. The results from this thesis can be used to reduce immunostimulation by any RNA when immune stimulation is not intended.  
date: 2014
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserver/17716/1/Dissertation%20Katharina%20Rimbach%2009%202014.pdf
identifier: DOI:10.11588/heidok.00017716
identifier: urn:nbn:de:bsz:16-heidok-177169
identifier:   Rimbach, Katharina   (2014) Inhibition of innate immune activation by modifications in bacterial RNA.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/17716/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng