Circularly permuted tRNA genes: their expression and implications for their physiological relevance and development

Abstract

A number of genome analyses and searches using programs that focus on the RNA-specific bulge-helix-bulge (BHB) motif have uncovered a wide variety of disrupted tRNA genes. The results of these analyses have shown that genetic information encoding functional RNAs is described in the genome cryptically and is retrieved using various strategies. One such strategy is represented by circularly permuted tRNA genes, in which the sequences encoding the 5'-half and 3'-half of the specific tRNA are separated and inverted on the genome. Biochemical analyses have defined a processing pathway in which the termini of tRNA precursors (pre-tRNAs) are ligated to form a characteristic circular RNA intermediate, which is then cleaved at the acceptor-stem to generate the typical cloverleaf structure with functional termini. The sequences adjacent to the processing site located between the 3'-half and the 5'-half of pre-tRNAs potentially form a BHB motif, which is the dominant recognition site for the tRNA-intron splicing endonuclease, suggesting that circularization of pre-tRNAs depends on the splicing machinery. Some permuted tRNAs contain a BHB-mediated intron in their 5'- or 3'-half, meaning that removal of an intron, as well as swapping of the 5'- and 3'-halves, are required during maturation of their pre-tRNAs. To date, 34 permuted tRNA genes have been identified from six species of unicellular algae and one archaeon. Although their physiological significance and mechanism of development remain unclear, the splicing system of BHB motifs seems to have played a key role in the formation of permuted tRNA genes. In this review, current knowledge of circularly permuted tRNA genes is presented and some unanswered questions regarding these species are discussed.

Keywords: BHB motif; circular gene permutation; intron; tRNA gene; tRNA-splicing endonuclease.

Figure 1

Gene organization and structures of permuted tRNAs. (A) Schematic representations of the structures of permuted tRNA genes with or without an intron. The 5′-half (blue) and the 3′-half (red) of the mature tRNA, the intron sequence (black), and the intervening sequence (green) of the pre-tRNA are shown. (B) Most permuted tRNAs can be classified into four types based on the location of the junction between the 3′-end of the 5′-half (blue) and the 5′-end of the 3′-half (red) in the secondary structure. (C) Inferred secondary structures of pre-tRNAs representing the four types of permuted tRNA genes in C. merolae. The arrowheads indicate the positions to be processed. The intron sequence is shown in lower case. The tRNA positions are numbered according to Marck and Grosjean (2002). The figures are partially identical to the Figure 1 of Soma et al. (2007).

Figure 2

Distribution of BHB motifs at the junctions of permuted tRNAs in algae and archaea. (A) The tRNA nucleotides are numbered according to Marck and Grosjean (2002) and the arrows indicate the positions of the BHB motifs. The type of BHB motif and the presence or absence of an intron is indicated for each tRNA. For example, “C. merolae Leu(TAA), HBh', +intron, (37/38)” at the position between nucleotides 20 and 21 means that the HBh' is from the junction of the C. merolae permuted tRNA^Leu(UAA), which contains an intron at 37/38. (B) The BHB motif is classified by one or two 3-nucleotide (nt) bulges (denoted as “B = 3”) separated by a central 4-base pair (bp) helix (denoted as “H = 4”) and flanked by two helices (denoted as h or h′), each with more than two base pairings. “hBHBh′” is the strict form of the BHB motif, which contains two bulges, a central helix and two flanking helices. The relaxed form of the BHB motif (BHL) lacking some of these bulges or helices is denoted as “hBH” or “HBh'.” The term “no H” represents motifs that do not contain a central 4-bp helix.

Figure 3

Comparison of the processing pathways for typical intronic and permuted pre-tRNAs. (A) Maturation of a typical intronic pre-tRNA involves intronic splicing, processing of the 5′- and 3′-ends by RNase P and tRNAse Z, and addition of the 3′-terminal CCA sequence. (B) Maturation of a permuted pre-tRNA starts with processing of the BHB motif (boxed) by the tRNA-splicing machinery, resulting in the formation of a circular RNA intermediate. The intervening sequence is then removed by RNase P and tRNase Z, followed by CCA addition. The sequential processing of permuted pre-tRNAs in the proposed pathway may be accomplished using processing machineries that are commonly used for typical pre-tRNAs, because the recognition elements for each processing enzyme are conserved in the permuted pre-tRNA and circular intermediate. The figure is partially identical to the Figure 3C of Soma et al. (2007).

Figure 4

Schematic representations of the tRNA-splicing endonuclease in archaea and eukaryotes. (A) An archaeal dimeric endonuclease comprising two catalytic subunits. (B) The yeast heterotetrameric endonuclease comprising two catalytic subunits (Sen2 and Sen34) and two accessory subunits (Sen15 and Sen54). (C) The possible heterodimeric (cmSen2 and cmSen34), heterotrimeric (cmSen2, cmSen34, and cmSen54), and heterotetrameric (cmSen2, cmSen34, cmSen54, and unidentified cmSen15) forms of the C. merolae endonuclease.

Figure 5

Proposed models for the development of permuted tRNA genes. (A) Permuted tRNA genes might have formed by gene duplication and loss of the outer segments. A tandem repeat of an intron-containing tRNA gene could be arranged into a permuted tRNA gene by the accumulation of mutations at both ends. Alternative models are based on reverse transcription of permuted or circularized pre-tRNA molecules. (B) A permuted pre-tRNA formed by an interaction between the 3′-half of the tRNA encoded by the initial gene and the 5′-half of the duplicated tRNA derived from the tandemly-repeated gene may have been reverse transcribed from the point indicated by an arrowhead and integrated back into the genome. (C) A circular intron-containing pre-tRNA produced by ligation of an intron-containing pre-tRNA at the acceptor-stem may also have been reverse transcribed and integrated into the genome.