Handling tRNA introns, archaeal way and eukaryotic way

Abstract

Introns are found in various tRNA genes in all the three kingdoms of life. Especially, archaeal and eukaryotic genomes are good sources of tRNA introns that are removed by proteinaceous splicing machinery. Most intron-containing tRNA genes both in archaea and eukaryotes possess an intron at a so-called canonical position, one nucleotide 3' to their anticodon, while recent bioinformatics have revealed unusual types of tRNA introns and their derivatives especially in archaeal genomes. Gain and loss of tRNA introns during various stages of evolution are obvious both in archaea and eukaryotes from analyses of comparative genomics. The splicing of tRNA molecules has been studied extensively from biochemical and cell biological points of view, and such analyses of eukaryotic systems provided interesting findings in the past years. Here, I summarize recent progresses in the analyses of tRNA introns and the splicing process, and try to clarify new and old questions to be solved in the next stages.

Keywords: archaea; eukaryote; genome; intron; splicing; tRNA.

Figure 1

Secondary structures of pre-tRNA and mature tRNA. Typical secondary structures of pre-tRNAs with a canonical intron (left) and mature tRNA (right) are schematically shown. White circles with three black circles, blues circles, and light gray circles represent 5′-exon, intron and 3′ exon of pre-tRNA, respectively. The anticodon is represented with black circles, and CCA tri-nucleotides added at the 3′-terminus of tRNA are depicted with dark gray circles. The BHB motif in archaeal pre-tRNA and the A-I pair in eukaryotic pre-tRNA are highlighted with orange and yellow shadow, respectively. Arrowheads represent splice sites. Mature tRNA is illustrated in two different ways. In the right most structure, decoding and amino acid-accepting units of mature tRNA are marked with light blue and light green shadow, respectively.

Figure 2

Insertion points of non-canonical introns in tRNA genes. Insertion points of non-canonical introns in various tRNA genes from archaea (A) and from eukaryotes (B) are shown. Positions of intron insertion found in certain groups of organisms are color-coded as shown on top of the tRNA models. Intron insertion points are summarized from data in Sugahara et al. (2008), Sugahara et al. (2009), and Chan et al. (2011) for archaea, and Kawach et al. (2005), Soma et al. (2007), and Maruyama et al. (2010) for eukaryotes.

Figure 3

Unusual tRNA genes. Various examples of unusual tRNA genes are represented (A–F). Exonic regions, BHB motifs and splice sites are marked as in Figure 1. Introns, leader and trailer sequences removed by splicing machinery are marked with bluish or purple-like colors. In panels (E,F), linker regions between permutated tRNA fragments are colored in green, and cleavage sites are marked with arrows. tRNA secondary structures are drawn according to Chan et al. (2011), Sugahara et al. (2007), Randau et al. (2005a), Fujishima et al. (2009), and Soma et al. (2007).

Figure 4

tRNA splice site cleavage and two pathways of tRNA ligation. Series of chemical reactions in splice site cleavage of pre-tRNA (left), the 5′-phosphate ligation pathway (upper), and the 3′-phosphate ligation pathway (lower) are shown schematically. Proteins involved in individual reactions are also shown, and unidentified factors are denoted by question marks. Those proteins of yeast, mammal and prokaryotes are color-coded with black, blue and orange, respectively. In the 5′-phosphate ligation pathway, yeast Tr11 uses GTP as a phosphate donor while hClp1 in the mammalian system uses ATP instead of GTP for 5′-phosphorylation of the 3′-exon. Although a mammalian enzyme(s) catalyzing the last two steps of this pathway is still missing, such an enzyme was found in lancelet. In the 3′-phosphate ligation pathway, the mammalian protein responsible for 2′-3′ cyclic phosphodiesterase in the HSPC117 complex was not fully identified, but eubacterial RtcB, an HSPC117 homolog, was demonstrated to possess this activity (Tanaka and Shuman, 2011). The starting material, pre-tRNA, and the end product, mature tRNA, are shadowed. Appr>Pi represents ADP-ribose 1″-2″ cyclic phosphate.

Figure 5

tRNA introns affect various biological activities as RNA and DNA units. Several examples where tRNA introns affect various biological activities as RNA units (upper) and as DNA units (lower) are illustrated. (A) The intron of tRNA-Ile_UAU in S. cerevisiae is an essential recognition motif for Pus1. Thus, pseudouridylation of U34 and U36 in the anticodon by Pus1 is only applied to the intron-containing pre-tRNA. Pre-tRNA-Ile_UAU pseudouridylated by Pus1 in the nucleus is subjected to cytoplasmic splicing after export from the nucleus in the yeast. (B) Yeast Trm5, a nuclear methyltransferase catalyzing the initial step of yW formation, only recognizes a spliced form of tRNA-Phe_GAA, which is produced in the cytoplasm and re-imported into the nucleus. Thus, yW formation starts at a late stage of tRNA-Phe_GAA maturation. (C) The intron of tRNA-Trp_CCA in H. volcanii is used as a guide RNA to select two nucleotides, C34 and U39, of the same tRNA for 2′-O-methylation. (D) Sulfolobus SSVs use a part of tRNAs as an attB site for its integration into the host genome. Some Sulfolobus species harbor an intron in such tRNA genes to disrupt attB sequences. This may be a bacterial strategy to avoid viral infection. (E) tRNA-Thr_AGU near the HMR-E locus, which is in a heterochromatic region, acts as an insulator and prevents expansion of the heterochromatic region over the tRNA gene by recruiting TFIIIC properly in S. cerevisiae. The intron-containing tRNA-Leu_CAA gene cannot replace tRNA-Thr_AGU while the intronless version of the same tRNA can. See the text for details.

Figure 6

Distribution of intron-containing tRNA genes in yeast genomes. Distribution of intron-containing genes in 6 representative yeast genomes (Saccharomyces cerevisiae, Candida glabrata, Lachancea thermotolerans, Kluyveromyces lactis, Eremothecium gossypii, and Debaryomyces hansenii) is summarized in the anti-codon table (A). As illustrated in the right, existence of intron-containing (orange) or intronless (blue) tRNA genes for each anticodon position is color-coded. Anticodon positions to which no tRNA genes are assigned on the genome or those correspond to termination codons are shown in white and black, respectively. The phylogenetic tree of these yeasts shown in (B) was drawn according to Génolevures Consortium (2009). Among these yeast, D. hansenii does not belong to Saccharomycetaceae and is used as an outgroup. (C) Sequence comparison of tRNA-Gly_UCC from D. hansenii (Dha), S. cerevisiae (Sce), C. glabrata (Cgl), L. thermotolerans (Lth), E. gossypii (Ego), and K. lactis (Kla), and tRNA-Glu_UUC from Lactobacillus plantarum (Lpl) and Clostridium tetanus (Cte). Nucleotides identical to those of K. lactis tRNA-Gly_UCC (marked with a frame) are shown in blue, and anticodons are highlighted with orange shadow.