<> "The repository administrator has not yet configured an RDF license."^^ . <> . . "Impact of nucleotide modifications on immune stimulatory effects of RNA"^^ . "Posttranscriptional RNA modifications are an important feature of self/non-self discrimination of nucleic acids by the innate immune system. Ribose 2’-O-methylation within RNA has been shown to limit recognition by Toll-like receptors (TLR) 7 and TLR8. In the natural RNA context, 2’-O-methylation of guanosine at position 18 (Gm18) within transfer RNAs (tRNAs) was identified to abolish RNA induced immune activation. However, the exact requirement of Gm18 within tRNA for affecting TLR responses remained unknown. Moreover, whether Gm18 plays a role as immune modulatory RNA modification in physiological settings or has different roles in pro- vs. eukaryotes was unexplored. Finally, preliminary data suggested the existence of further RNA modifications that affect immune recognition. This thesis therefore aimed to explore the function of Gm18 for immune regulation in more detail and to study further RNA modifications for immune regulatory properties.\r\nIn a combined chemical-immunological approach on human tRNALys3 which showed no TLR7 activation but lacked Gm18, this work identified a new immune-silencing modification: Tm, a double-methylation of uridine, at position 54 of human tRNAs was sufficient to decrease immune stimulation of TLR7. However, in contrast to Gm18, the effects were not dominant negative in co-stimulation experiments. Thus, for the first time an immune silencing modification in a natural RNA context has been deciphered.\r\nTo further define the inhibition of TLR7 and TLR8 activation by Gm18 modified RNA, a systematic variation of nucleotides within a conserved tRNA sequence was performed. The studies allowed identification of an optimized sequence motif antagonizing TLR7/8 responses. The minimal, naturally occurring GmGC tri-nucleotide motif within a 9-mer oligoribonucleotide was identified as most efficient to antagonize recognition of otherwise immune stimulatory RNA. The findings could be beneficial to design immune inhibitory oligoribonucleotides as therapeutic approach in autoimmune diseases associated with exaggerated TLR7/8 activation. \r\nIn further series of experiments mutants lacking Gm18 modification in either bacterial or eukaryotic tRNA were used to study the functional impact on overall immune stimulation. tRNA from an Escherichia coli (E.coli) mutant lacking the enzyme trmH that incorporates Gm18 showed increased immune stimulation. However, stimulation of human peripheral blood mononuclear cells (PBMCs) and TLR8 deficient genetically engineered monocyte/macrophage–like BlaER1 cells, with whole E.coli wild-type and trmH deficient bacteria revealed RNA dependent recognition of E. coli but negligible effects of Gm18. Interestingly, results indicate that trmH mediated Gm18 modification might be subject to regulation under stress conditions. Similarly, CRISPR/Cas9 generated knockouts of TARBP1, the Gm18 methyltransferase in human cells, showed slightly increased immunostimulation for tRNA fractions but no change for whole RNA stimulation. \r\nIn summary, RNA 2’-O-methyl modification within tRNA is capable to limit immune recognition by TLR7/8, yet for manipulation of the overall immune recognition of bacterial or eukaryotic RNA the identified modifications alone seem to be insufficient."^^ . "2018" . . . . . . . "Isabel"^^ . "Freund"^^ . "Isabel Freund"^^ . . . . . . "Impact of nucleotide modifications on immune stimulatory effects of RNA (PDF)"^^ . . . "MyOutPutPDF-A.pdf"^^ . . . "Impact of nucleotide modifications on immune stimulatory effects of RNA (Other)"^^ . . . . . . "indexcodes.txt"^^ . . "HTML Summary of #25688 \n\nImpact of nucleotide modifications on immune stimulatory effects of RNA\n\n" . "text/html" . . . "570 Biowissenschaften, Biologie"@de . "570 Life sciences"@en . .