Cutting, dicing, healing and sealing: the molecular surgery of tRNA

Abstract

All organisms encode transfer RNAs (tRNAs) that are synthesized as precursor molecules bearing extra sequences at their 5' and 3' ends; some tRNAs also contain introns, which are removed by splicing. Despite commonality in what the ultimate goal is (i.e., producing a mature tRNA), mechanistically, tRNA splicing differs between Bacteria and Archaea or Eukarya. The number and position of tRNA introns varies between organisms and even between different tRNAs within the same organism, suggesting a degree of plasticity in both the evolution and persistence of modern tRNA splicing systems. Here we will review recent findings that not only highlight nuances in splicing pathways but also provide potential reasons for the maintenance of introns in tRNA. Recently, connections between defects in the components of the tRNA splicing machinery and medically relevant phenotypes in humans have been reported. These differences will be discussed in terms of the importance of splicing for tRNA function and in a broader context on how tRNA splicing defects can often have unpredictable consequences.

FIGURE 1

The substrates of tRNA-splicing endonucleases of Archaea and Eukarya. Dashed lines represent the splicing determinants taken from eukaryotic and archaeal tRNA to create tRNA^ArchEuka. tRNA^ArchEuka is composed of the body of a eukaryotic tRNA (top half) with the anticodon stem of an Archaeal tRNA (box). The box identifies the canonical bulge-helix-bulge (BHB) motif of Archaeal endonucleases, while the eukaryotic cardinal positions and the A–I interaction are denoted in yellow.

FIGURE 2

The two main tRNA-splicing pathways. The 5^′→3^′ ligation pathway (top) has been extensively studied in yeast. Following cleavage by the tRNA-splicing endonuclease, the resulting exon halves are processed by the CPD and kinase domains; the CPD domains generates a 3^′ terminus 2^′-phospate on the 5^′ exon and the kinase domain phosporylates the 5^′ end of the 3^′ exon using GTP. The two exons are joined by the ligase domain of the same protein (Trl1) using ATP, the dangling 2^′-phosphate is finally removed by Tpt1 which is a free-standing enzyme. All domains reside in a single Trl1 protein and are shown in yellow boxes. The 3^′→5^′ ligation pathway has been described in archaea and vertebrates. RtcB (green box) is the enzyme responsible for the opening of the 2^′3^′ cyclic phosphate (in some organisms but not in mammals) and for the joining of the halves. In humans RtcB named HSPC117 has been identified as part of the human ligase complex.

FIGURE 3

Subcellular localization of the tRNA-splicing machinery. In yeast (in pink), the endonuclease complex is localized on the outer membrane surface of mitochondria. The precursor tRNA is cleaved and the halves are processed and joined by Trl1 in the cytoplasm. The remaining 2^′-phosphate is removed by Tpt1, which is also localized in the cytoplasm, generating the spliced tRNA. In humans, splicing is accomplished by a nuclear macromolecular complex formed by the endonuclease (pink), the Clp1 kinase (green), HSPC117/RtcB (light blue), and other proteins (purple).