The Functions of Chloroplast Glutamyl-tRNA in Translation and Tetrapyrrole Biosynthesis1[OPEN]

The generation and characterization of transplastomic tobacco plants provide insight into the dual role of the chloroplast glutamyl-tRNA in protein biosynthesis and tetrapyrrole biosynthesis. The chloroplast glutamyl-tRNA (tRNA(Glu)) is unique in that it has two entirely different functions. In addition to acting in translation, it serves as the substrate of glutamyl-tRNA reductase (GluTR), the enzyme catalyzing the committed step in the tetrapyrrole biosynthetic pathway. How the tRNA(Glu) pool is distributed between the two pathways and whether tRNA(Glu) allocation limits tetrapyrrole biosynthesis and/or protein biosynthesis remains poorly understood. We generated a series of transplastomic tobacco (Nicotiana tabacum) plants to alter tRNA(Glu) expression levels and introduced a point mutation into the plastid trnE gene, which has been reported to uncouple protein biosynthesis from tetrapyrrole biosynthesis in chloroplasts of the protist Euglena gracilis. We show that, rather than comparable uncoupling of the two pathways, the trnE mutation is lethal in tobacco because it inhibits tRNA processing, thus preventing translation of Glu codons. Ectopic expression of the mutated trnE gene uncovered an unexpected inhibition of glutamyl-tRNA reductase by immature tRNA(Glu). We further demonstrate that whereas overexpression of tRNA(Glu) does not affect tetrapyrrole biosynthesis, reduction of GluTR activity through inhibition by tRNA(Glu) precursors causes tetrapyrrole synthesis to become limiting in early plant development when active photosystem biogenesis provokes a high demand for de novo chlorophyll biosynthesis. Taken together, our findings provide insight into the roles of tRNA(Glu) at the intersection of protein biosynthesis and tetrapyrrole biosynthesis.