The chloroplast atpA transcript in Chlamydomonas reinhardtii is specifically activated for translation by the nucleus-encoded TDA1 protein or stored, when not translated, in non-polysomic ribonucleoprotein complexes
Stephan Eberhard, Christelle Loiselay, Dominique Drapier, Sandrine Bujaldon, Richard Kuras, Yves Choquet, and Francis-André Wollman
UMR 7141 (CNRS - Université Paris 6) Institut de Biologie Physico-Chimique 13, rue Pierre et Marie Curie F-75005 Paris, France
We cloned the nucleus-encoded TDA1 gene specifically required for translation of the chloroplast atpA transcript. This 1267 aa protein contains a predicted chloroplast-targeting sequence and octopentatricopeptid (OPR) repeats in its C-terminus. This domain ressembles double-helical PPR domains involved in Protein/RNA interactions, found in nucleus-encoded regulatory proteins acting on organellar gene expression. Using chimeric chloroplast genes and truncated versions within the TDA1 coding region, we show (i) that the molecular target of TDA1 lies in the 5' UTR of the atpA transcript and (ii) that the 667 aa C-terminal part of TDA1 containing the OPR motifs is sufficient to complement a tda1 mutant strain. We also investigated the fate of non-translated atpA mRNAs, since we have shown previously that only a small fraction of atpA mRNAs are directed to translation at any given time (Eberhard et al. 2002). To this end, we compared the distribution of atpA transcripts on sucrose gradients in the WT and in mutant strains defective for atpA translation. In both a nuclear mutant in the TDA1 gene and in another mutant strain bearing a chloroplast mutation in the atpA gene that converts the initiation codon into a stop-codon, non-translated atpA transcripts are detected in high density fractions of the sucrose gradient. Using chimeric genes, we show that the 5' UTR of the atpA transcript is sufficient to recruit unrelated coding sequences to these non polysomic complexes. Our results suggest that the 5' UTR of the atpA transcript is thus involved (i) in its association with polysomes, in a TDA1-dependent manner, and (ii) in its storage in non-polysomic complexes when it can not be translated. We discuss the physiological significance of this balance between translational activation and storage of chloroplast transcripts.
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