Post-transcriptional modifications to tRNA--a response to the genetic code degeneracy.

We used to think of the genetic code degeneracy as an obligatory process resulting from assigning 61 sense codons to the 20 amino acids in protein. In this degeneracy, 18 of the 20 amino acids are endowed with multiple codons, and the conventional wisdom was that each of the 18 was translated equally from each codon. However, due to the degeneracy mostly manifest at the wobble position for each amino acid, we were led to believe that the degeneracy is simply to provide a deck of codons for safety control, in case one or two codons are damaged by nucleotide substitutions to the wobble position. However, work over a long path has now shown that the degeneracy has a much deeper meaning, with the ability to confer codon-by-codon translational regulation. At its core, translation of a codon requires pairing interaction with the anticodon of its cognate tRNA. Due to the degeneracy, multiple codons are paired with one anticodon in different efficiencies and stabilities. This is a form of translational regulation executed at the level of individual codons. Nature has developed this translational regulation by two means: one by employing biased codon usage and the other by modifying the chemical structure of the tRNA anticodon or adjacent nucleotides via post-transcriptional modifications. While biased codon usage is genetically encoded, post-transcriptional modifications are epigenetic and have more flexibility to modulate the fidelity and efficiency of codon-anticodon pairing interactions. Indeed, among codons for the same amino acid, some require tRNA modifications that are stress-induced, conferring a capacity to regulate cellular adaptation to stress at individual codons. Thus, in response to the genetic code degeneracy, tRNA has evolved to carry post-transcriptional modifications to fine-tune translation one codon at a time. Enzymes catalyzing tRNA modifications are often linked to human diseases, ranging from metabolic defects, mitochondrial dysfunctions, and neurological disorders, to cancer. The greatest diversity of tRNAmodifications is at positions 34 (the wobble position) and 37 (on the 3′ side of the anticodon), two positions critical for translational accuracy and reading-frame maintenance. Three modifications at these positions have strong effects on codon-anticodon pairing interactions. First, U34 is modified to 5-methoxycarbonyl-methyl-2-thiouridine (mcmsU34) in tRNA (anticodon 5′-UUU-3′), tRNA, tRNA, and tRNA. In the case of tRNA for example, while it can read both Lys codons (5′-AAA and 5′-AAG), the codon-anticodon pairing in either case is weak, consisting of A-U and G-U base pairs. The addition of mcms to U34 can stabilize the A-U and G-U pairs at the wobble position by providing an extended chemical group to enhance the stacking interaction within each pair. The importance of this mcms modification is emphasized by its response to cellular stress. Specifically, mcmsU34 is synthesized in two sequential steps, first via the addition of mcm to U34 by the ELP complex and second via the addition of s to the mcmU34 intermediate by the URM1 pathway. While the ELP complex is formally associated with the RNA pol II transcription complex in yeast, it catalyzes themcm addition to U34 upon cellular oxidative stress. Similarly, the URM1 complex is activated by cellular stress in yeast, such as oxidative stress and temperature up-shift stress. Recent work shows that mcms addition to U34 facilitates translation of codonbiased mRNAs required for stress response. For example, upon stress with the alkylating agent methyl methane sulfonate, the methyl transferase Trm9 associated with the ELP complex is activated to help to translate DNA damage response genes each with a pattern of codon usage dependent on mcmsU34-tRNA for translation. Similarly, a human Trm9-like protein in cancer cells helps to translate the LIN9 protein, which has a codon bias requiring mcmsU34tRNA for translation. LIN9 interacts with the tumor suppressor protein Rb to suppress cancer progression, thus implicating the Trm9-like protein and the mcms modification of U34-tRNA with a role in inhibition of tumor growth.