Anti-transfer RNA antibodies in two patients with pulmonary fibrosis, Raynaud's phenomenon and polyarthritis

Abstract

Patient W.S. (a 61-year-old woman) and patient T.M. (a 41-year-old man) developed recurrent fevers, polyarthritis, Raynaud's phenomenon and interstitial pulmonary fibrosis without apparent polymyositis. From HeLa cell extracts, sera from both patients immunoprecipitated all species of intact and deproteinised tRNAs. To identify the antibody binding site more precisely, tRNAs transcribed in vitro from cloned Escherichia coli tRNA genes and various mutants were prepared and used as antigens for immunoprecipitation. When the TpsiC loop, or the D loop were deleted, such mutants were not bound by both sera, suggesting that the D and TpsiC loops were required for antibody binding. Abrogation of tRNA binding occurred when 18G of tRNATrp was replaced with 18A to break the tertiary L-shape structure of tRNA. These results strongly suggest that sera from W.S. and T.M. recognise the tertiary conformation of L-shaped tRNA which is constructed with both D and TpsiC loops. These autoantibodies may also serve as a marker for a new subset of patients with connective tissue diseases that is distinct from anti-aminoacyl-tRNA synthetase syndrome.

Synthetic antibodies for specific recognition and crystallization of structured RNA

“…A large number of these RNAs adopt complex three-dimensional architectures that frequently act in complex with proteins to mediate their biological function (12,13). Nevertheless, with the exception of a handful of examples, mostly isolated from the sera of autoimmune patients (14)(15)(16)(17), we know little about anti-RNA antibodies and their recognition of nucleic acids. This dearth of information reflects our inability to elicit antibodies against RNA by using traditional approaches.…”

Inosine and N 1-methylinosine within a synthetic oligomer mimicking the anticodon loop of human tRNAAla are major epitopes for anti-PL-12 myositis autoantibodies

“…Several autoantibodies that recognize various deproteinized RNAs have been described (reviewed in Hoet & Van Venrooij, 1992;Keene, 1996;Mimori, 1996)+ Among them, only those against U1-U5 snRNA, as detected in few patients with systemic sclerosis, were shown to be directed to a specific modified nucleoside (2,2,7-trimethylguanosine cap structure) (Okano & -methylinosinic acid (pm 1 I) are higher in F than in E because the ratio between modified and nonmodified nucleotides in the variant RNA 7 is higher than for variant RNA 5+ The solvent system 1 as described in Auxilien et al+ (1996) was used in all cases+ Medsger, 1992)+ Except for the autoantibodies against tRNA Ala (anticodon IGC) described in the present study, the other anti-RNA autoantibodies characterized so far are not directed to modified nucleotides, as they also immunoprecipitate in vitro transcripts, lacking modified nucleosides, of the natural or synthetic genes corresponding to these antigenic RNAs+ Examples include autoantibodies against a characteristic 40-nt stem-loop fragment in U1snRNA (Deutscher & Keene, 1988;Tsai et al+, 1992;Teunissen et al+, 1998), a 59-nt fragment of 28S rRNA (Chu et al+, 1991), a 84-nt fragment of Y5 Ro RNA (Boulanger et al+, 1995), or various mutants of tRNA retaining their L-shaped three-dimensional architecture (Matsumura et al+, 1996;Brouwer et al+, 1998;Ohosone et al+, 1998)+ In these cases a few selected canonical nucleotides within a well-defined RNA conformation (a characteristic stem-loop or a three-dimensional architecture) constitute the major epitopes recognized by the corresponding IgG+ It is worth mentioning that antibodies predominantly directed towards the characteristic hypermodified nucleoside wybutosine, which is located at position 37 of the anticodon loop of yeast tRNA Phe , were obtained by immunizing a goat with a glutaraldehyde-mediated conjugate of yeast tRNA Phe and bovine gamma globulin (Fuchs et al+, 1974)+ Also, antibodies against a variety of modified nucleosides, including inosine (Inouye et al+, 1971) and 1-methylinosine (D'Ambrosio et al+, 1991; Renaud et al+, 1991) have been prepared by immunizing rabbits, mice or goats with a conjugate of the hapten and a protein carrier (reviewed in Vold, 1990)+ In this article, we present evidence that the important features recognized by human anti-tRNA Ala (anti-PL-12) autoantibodies include the chemical identity and the spatial disposition of two modified ribonucleotides within the anticodon loop of tRNA Ala (anticodon IGC): namely the bases inosine at position 34 and N 1 -methylinosine at position 37 (see Fig+ 1A), their spacing, and probably their three-dimensional configuration+ Figure 5 shows the spatial orientation of the 6-keto groups (in black) of inosine-34 and of the N 1 -methyl group (in grey) of N 1 -methylinosi...…”

Crystal structure of an RNA polymerase ribozyme in complex with an antibody fragment

“…In contrast, we know very little about antibodies that bind to RNA, other than the few isolated from the sera of autoimmune patients [29][30][31][32]. For example, antibodies that bind to the U1 small nuclear RNA (and many mammalian small RNA -protein complexes) have been isolated from patients with autoimmune diseases [33].…”