Distinct localization of chiral proofreaders resolves organellar translation conflict in plants.

Plants have two endosymbiotic organelles originated from two bacterial ancestors. The transition from an independent bacterium to a successful organelle would have required extensive rewiring of biochemical networks for its integration with archaeal host. Here, using Arabidopsis as a model system, we show that plant D-aminoacyl-tRNA deacylase 1 (DTD1), of bacterial origin, is detrimental to organellar protein synthesis owing to its changed tRNA recognition code. Plants survive this conflict by spatially restricting the conflicted DTD1 to the cytosol. In addition, plants have targeted archaeal DTD2 to both the organelles as it is compatible with their translation machinery due to its strict D-chiral specificity and lack of tRNA determinants. Intriguingly, plants have confined bacterial-derived DTD1 to work in archaeal-derived cytosolic compartment whereas archaeal DTD2 is targeted to bacterial-derived organelles. Overall, the study provides a remarkable example of the criticality of optimization of biochemical networks for survival and evolution of plant mitochondria and chloroplast.

Switching a conflicted bacterial DTD-tRNA code is essential for the emergence of mitochondria.

Mitochondria emerged through an endosymbiotic event involving a proteobacterium and an archaeal host. However, the process of optimization of cellular processes required for the successful evolution and survival of mitochondria, which integrates components from two evolutionarily distinct ancestors as well as novel eukaryotic elements, is not well understood. We identify two key switches in the translational machinery­one in the discriminator recognition code of a chiral proofreader DTD [d-aminoacyl­transfer RNA (tRNA) deacylase] and the other in mitochondrial tRNAGly­that enable the compatibility between disparate elements essential for survival. Notably, the mito-tRNAGly discriminator element is the only one to switch from pyrimidine to purine during the bacteria-to-mitochondria transition. We capture this code transition in the Jakobida, an early diverging eukaryotic clade bearing the most bacterial-like mito-genome, wherein both discriminator elements are present. This study underscores the need to explore the fundamental integration strategies critical for mitochondrial and eukaryotic evolution.

Recruitment of archaeal DTD is a key event toward the emergence of land plants.

Streptophyte algae emerged as a land plant with adaptations that eventually led to terrestrialization. Land plants encounter a range of biotic and abiotic stresses that elicit anaerobic stress responses. Here, we show that acetaldehyde, a toxic metabolite of anaerobic stress, targets and generates ethyl adducts on aminoacyl-tRNA, a central component of the translation machinery. However, elongation factor thermo unstable (EF-Tu) safeguards l-aminoacyl-tRNA, but not d-aminoacyl-tRNA, from being modified by acetaldehyde. We identified a unique activity of archaeal-derived chiral proofreading module, d-aminoacyl-tRNA deacylase 2 (DTD2), that removes N-ethyl adducts formed on d-aminoacyl-tRNAs (NEDATs). Thus, the study provides the molecular basis of ethanol and acetaldehyde hypersensitivity in DTD2 knockout plants. We uncovered an important gene transfer event from methanogenic archaea to the ancestor of land plants. While missing in other algal lineages, DTD2 is conserved from streptophyte algae to land plants, suggesting its role toward the emergence and evolution of land plants.

A chiral selectivity relaxed paralog of DTD for proofreading tRNA mischarging in Animalia.

D-aminoacyl-tRNA deacylase (DTD), a bacterial/eukaryotic trans-editing factor, removes D-amino acids mischarged on tRNAs and achiral glycine mischarged on tRNAAla. An invariant cross-subunit Gly-cisPro motif forms the mechanistic basis of L-amino acid rejection from the catalytic site. Here, we present the identification of a DTD variant, named ATD (Animalia-specific tRNA deacylase), that harbors a Gly-transPro motif. The cis-to-trans switch causes a "gain of function" through L-chiral selectivity in ATD resulting in the clearing of L-alanine mischarged on tRNAThr(G4•U69) by eukaryotic AlaRS. The proofreading activity of ATD is conserved across diverse classes of phylum Chordata. Animalia genomes enriched in tRNAThr(G4•U69) genes are in strict association with the presence of ATD, underlining the mandatory requirement of a dedicated factor to proofread tRNA misaminoacylation. The study highlights the emergence of ATD during genome expansion as a key event associated with the evolution of Animalia.