Natural reassignment of CUU and CUA sense codons to alanine in Ashbya mitochondria.
The discovery of diverse codon reassignment events has demonstrated that the canonical genetic code is not universal. Studying coding reassignment at the molecular level is critical for understanding genetic code evolution, and provides clues to genetic code manipulation in synthetic biology. Here we report a novel reassignment event in the mitochondria of Ashbya (Eremothecium) gossypii, a filamentous-growing plant pathogen related to yeast (Saccharomycetaceae). Bioinformatics studies of conserved positions in mitochondrial DNA-encoded proteins suggest that CUU and CUA codons correspond to alanine in A. gossypii, instead of leucine in the standard code or threonine in yeast mitochondria. Reassignment of CUA to Ala was confirmed at the protein level by mass spectrometry. We further demonstrate that a predicted tRNA(Ala)UAG is transcribed and accurately processed in vivo, and is responsible for Ala reassignment. Enzymatic studies reveal that tRNA(Ala)UAG is efficiently recognized by A. gossypii mitochondrial alanyl-tRNA synthetase (AgAlaRS). AlaRS typically recognizes the G3:U70 base pair of tRNA(Ala); a G3A change in Ashbya tRNA(Ala)UAG abolishes its recognition by AgAlaRS. Conversely, an A3G mutation in Saccharomyces cerevisiae tRNA(Thr)UAG confers tRNA recognition by AgAlaRS. Our work highlights the dynamic feature of natural genetic codes in mitochondria, and the relative simplicity by which tRNA identity may be switched.
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520-8114, USA, Département de Biochimie, Centre Robert-Cedergren, Université de Montréal, 2900 Boulevard Edouard Montpetit, Montréal, Québec, H3C 3J7, Canada, Program in Chemical Biology, Harvard University, Cambridge, MA 02138, USA, Department of Genetics, Harvard Medical School, Boston, MA 02115, USA and Department of Chemistry, Yale University, New Haven, CT 06520-8114, USA.
Jiqiang
J
Ling
Rachid
R
Daoud
Marc J
MJ
Lajoie
George M
GM
Church
Dieter
D
Söll
B Franz
BF
Lang
D000409:Alanine / Q000378:metabolism; D000412:Alanine-tRNA Ligase / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D003062:Codon; D055169:Eremothecium / Q000201:enzymology / Q000235:genetics*; D007930:Leucine / Q000378:metabolism; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism; D012344:RNA, Transfer, Ala / Q000737:chemistry / Q000378:metabolism*
D003062:Codon; D024101:Mitochondrial Proteins; D012344:RNA, Transfer, Ala; D012343:RNA, Transfer; D000412:Alanine-tRNA Ligase; D007930:Leucine; D000409:Alanine