Additional pathway to translate the downstream ndhK cistron in partially overlapping ndhC-ndhK mRNAs in chloroplasts

Abstract

The chloroplast NAD(P)H dehydrogenase (NDH) C (ndhC) and ndhK genes partially overlap and are cotranscribed in many plants. We previously reported that the tobacco ndhC/K genes are translationally coupled but produce NdhC and NdhK, subunits of the NDH complex, in similar amounts. Generally, translation of the downstreamcistron in overlapping mRNAs is very low. Hence, these findings suggested that the ndhK cistron is translated not only from the ndhC 5′UTR but also by an additional pathway. Using an in vitro translation system from tobacco chloroplasts, we report here that free ribosomes enter, with formylmethionyl-tRNAfMet, at an internal AUG start codon that is located in frame in the middle of the upstream ndhC cistron, translate the 3′ half of the ndhC cistron, reach the ndhK start codon, and that, at that point, some ribosomes resume ndhK translation. We detected a peptide corresponding to a 57-amino-acid product encoded by the sequence from the internal AUG to the ndhC stop codon. We propose a model in which the internal initiation site AUG is not designed for synthesizing a functional isoform but for delivering additional ribosomes to the ndhK cistron to produce NdhK in the amount required for the assembly of the NDH complex. This pathway is a unique type of translation to produce protein in the needed amount with the cost of peptide synthesis.