Emerging functional principles of tRNA-derived small RNAs and other regulatory small RNAs

Abstract

Recent advancements in small RNA sequencing have unveiled a previously hidden world of regulatory small noncoding RNAs (sncRNAs) that extend beyond the well-studied small interfering RNAs, microRNAs, and piwi-interacting RNAs. This exploration, starting with tRNA-derived small RNAs, has led to the discovery of a diverse universe of sncRNAs derived from various longer structured RNAs such as rRNAs, small nucleolar RNAs, small nuclear RNAs, Y RNAs, and vault RNAs, with exciting uncharted functional possibilities. In this perspective, we discuss the emerging functional principles of sncRNAs beyond the well-known RNAi-like mechanisms, focusing on those that operate independent of linear sequence complementarity but rather function in an aptamer-like fashion. Aptamers use 3D structure for specific interactions with ligands and are modulated by RNA modifications and subcellular environments. Given that aptamer-like sncRNA functions are widespread and present in species lacking RNAi, they may represent an ancient functional principle that predates RNAi. We propose a rethinking of the origin of RNAi and its relationship with these aptamer-like functions in sncRNAs and how these complementary mechanisms shape biological processes. Lastly, the aptamer-like function of sncRNAs highlights the need for caution in using small RNA mimics in research and therapeutics, as their specificity is not restricted solely to linear sequence.

Keywords: AGO; PANDORA-seq; RNA modification; RNA structure; Y RNA; aptamer; rRNA; small RNA; snRNA; snoRNA; tRNA; vault RNA.

Figure 1

Schematics of different sncRNAs derived from longer parental RNAs Ref. (1).A, tRNA-derived small RNA (tsRNA); (B) snRNA-derived small RNA (snsRNA); (C) rRNA-derived small RNA (rsRNA); (D) Y RNA–derived small RNA (ysRNA); (E) vault RNA–derived small RNA (vtsRNA); (F) snoRNA-derived small RNA (snosRNA). The upper section of each panel illustrates the mapping data of sncRNAs from their parental RNAs, with numbers in the y-axis indicating reads per million (RPM). The lower section of each panel presents the predicted secondary structure of each parental RNA (depicted in gray) according to RNAcentral (https://rnacentral.org). The regions from which sncRNAs are derived are highlighted in color, corresponding to the peaks in the upper section of the panel. sncRNA, small noncoding RNA.

Figure 2

miRNAs represent a minority of the total sncRNA repertoire.A, quantity of different sncRNAs present across various mouse tissues and cells. Data are derived from Ref. (12). B, sequence diversity of different sncRNAs in the mouse, including the cumulative sum of mouse miRNA types sourced from miRbase. The types of other detected sncRNAs are compiled from pooled data of tissues and cells provided in Ref. (12). The sncRNAs depicted in the inner circle represent detected sncRNAs that appear in at least two independent sequencing libraries. The outer circles represent the theoretical possibilities for sncRNA generation, based on potential cleavage at any position of the RNAs (i.e., Y RNAs, vtRNAs, tRNAs, rRNAs, snRNAs, and snoRNAs) to produce sncRNA products of 15 to 44 nucleotides in length. sncRNA, small noncoding RNA; snoRNA, small nucleolar RNA; snRNA, small nuclear RNA; vtRNA, vault RNA.

Figure 3

Enzymatic regulation of tRNA:tsRNA dynamics. Schematics of tRNA cleavage by RNases, followed by either tRNA repair or selective degradation of tRNA to generate specific tsRNAs. RNase, ribonuclease; tsRNA, tRNA-derived small RNA.

Figure 4

Predicted secondary and tertiary structures for individual sncRNA sequences. (A) miR-1, (B) tsRNA-Gly-GCC, and (C) rsRNA-28s. RNAstructure (177) and RNAComposer (178) were used to predict the secondary and tertiary structure of the sncRNAs, respectively. The forna tool (part of ViennaRNA) (179) and UCSF Chimera (180) were used to visualize the secondary and tertiary structure, respectively. The standard free energy difference (ΔG° value in kcal/mol) between the folded and unfolded forms of the RNA, predicted at 37 °C, is also presented. Notably, the positive ΔG° for miR-1 suggests a preference for unfolded state. Nevertheless, miR-1 has been reported to act as an aptamer by binding with the potassium channel Kir2.1 to change channel activity (135). This suggests that small RNAs can function as aptamers even through their linear form, further underscoring the versatility of aptamer-like functions for small RNAs. rsRNA, rRNA-derived small RNA; sncRNA, small noncoding RNA; tsRNA, tRNA-derived small RNA.

Figure 5

Different modes of sncRNA function, using linear base pairing, 3D structure-based aptamer function, and a combination of both.A, complementarity to target sequence without protein binding. B, protein-assisted RNAi-based linear base pairing. C, binding with protein using a 3D structure as an aptamer. Note that the sncRNA can fold either on its own or multimerize with other RNAs to form higher-order structures for binding activity. D, combination of linear base pairing and aptamer-like function with different domains. For a single sncRNA, the listed functional modes might be interchangeable under different subcellular environments (e.g., cytoplasmic, nuclear, and mitochondrial) and induced by specific local factors. RNP, ribonucleoprotein; sncRNA, small noncoding RNA.