MODIFIED NUCLEOSIDES OF SACCHAROMYCES CEREVISIAE RNA: ANALYTICAL PROFILING AND POTENTIAL RELEVANCE TO INNATE IMMUNE RECOGNITION

Abstract

Natural Saccharomyces cerevisiae RNAs are chemically and structurally heterogeneous polynucleotide systems. Their pharmaceutical-chemical charac-terization has traditionally relied on origin, total RNA content, molecular-size distribution, polymer integrity, secondary structure and impurity profile. However, cellular RNAs also contain numerous post-transcriptional nucleoside modifications that constitute an additional epitranscriptomic level of mo-lecular organization. The aim of this review was to systematize current evidence on modified nucleosides in S. cerevisiae, evaluate their potential rele-vance to endolysosomal processing and TLR7/8-dependent RNA sensing, and substantiate analytical approaches to epitranscriptomic profiling of natural yeast-RNA substances. Original studies and reviews indexed in PubMed/MEDLINE or available from scientific publishers were analyzed, with priority giv-en to 2021–2026 publications; earlier landmark studies were included when essential for established mechanisms or analytical methodology. Chemical heterogeneity of yeast RNA extends beyond chain-length distribution and polymer integrity. Mass-spectrometric studies have identified m7G, m6A, Gm, Cm, ac4C and f5C in S. cerevisiae mRNA, whereas tRNA and rRNA display an even richer modification landscape that can change under cellular stress. The strongest mechanistic evidence concerning innate immune sensing currently relates to pseudouridine and 2′-O-methylation, which can modify susceptibility to endolysosomal nucleases and thereby alter the generation of TLR7/8 ligands. These findings do not establish a causal relationship be-tween batch-to-batch epitranscriptomic variability of industrial yeast RNA and immunobiological activity, but they provide a testable mechanistic framework. A tiered analytical strategy is proposed that combines enzymatic RNA digestion, quantitative LC–MS/MS profiling of modified nucleosides using internal standards, fraction-specific analysis, control of sample-preparation artifacts, and correlation of the chemical fingerprint with functional in vitro assays. The epitranscriptomic profile should currently be regarded as a candidate molecular descriptor of natural yeast RNA; designation as a crit-ical quality attribute requires direct experimental validation.