Light regulates the rate of translation elongation of chloroplast reaction center protein D1

Abstract

Intact and lysed chloroplasts isolated from the day or night phase of seedling growth exhibit a higher rate of [ 35 S]Met incorporation into the D1 protein in the light than in darkness. In the presence of the translation initiation inhibitor lincomycin, radiolabel incorporation remains unaffected for 7.5Ϫ10 min of the in vitro translation reaction, indicating that radiolabel incorporation is regulated by translation elongation. The rate of [ 35 S]Met incorporation into D1-protein can be increased by addition of exogenous ATP to the in vitro translation reactions; however, ATP cannot replace light, and at physiological concentrations of stromal ATP (40 µM), the rate is at least 25-fold higher in the light than in darkness. This indicates that translation elongation is arrested in darkness. Separation of translation-elongation reactions into polysome-bound and membrane-integrated D1 proteins demonstrates that the rate of translation elongation is higher in the presence of light. In the light, less time is required to transiently radiolabel a D1 translation intermediate of about 17 kDa and to chase the translation intermediate into mature D1 protein.We propose that light regulates the enzymatic activity of the translation-elongation process in chloroplasts.Keywords : chloroplast; translation elongation ; D1.Posttranscriptional regulation of gene expression in higher plant plastids has been intensively studied [1,2]. Studies of gene expression during plastid development and in mature chloroplasts have provided evidence for a light-induced regulation of translation [3Ϫ6]. Specifically, the light-regulated translation of the psbA mRNA encoding the photosystem II reaction-center protein D1 has been investigated in detail.In etioplasts isolated from 4-day-old dark-grown barley seedlings, the psbA mRNA was shown to be continuously translated ; however, the D1 protein was degraded in the absence of chlorophyll (Chl) and stabilized by binding of de novo synthesized Chl [7Ϫ10]. When 7Ϫ8-day-old dark-grown barley was illuminated, accumulation of D1 was dependent on developmentally regulated cytoplasmic factors and the psbA mRNA was found to be regulated on the level of translation initiation [11Ϫ 14]. In the light, analysis of ribosome distribution on psbA mRNA supported an accumulation of ribosomes at the site of initiation, but also revealed that ribosomes pause during translation elongation [14Ϫ16]. Pausing of ribosomes was speculated to improve the efficiency of Chl binding to D1 nascent chains or to enable integration of the D1 protein into the etioplast membranes during greening [9,15].In plastids isolated from transgenic tobacco leaves and in Chlamydomonas reinhardtii, light-dependent accumulation of D1 has been ascribed to regulation at the 5′-untranslated region of the psbA mRNA [17Ϫ19] ; structured RNA elements were characterized to be responsible for translation initiation and proteins were identified that specifically bind to the 5′-untranslated region during translation initiation [17,20,21] nas, ADP-dependent ...

Chloroplast protein targeting involves localized translation in Chlamydomonas

“…To determine whether these rbcL mRNAs are localized by targeting signals in the LSU nascent chain (in a cotranslational mechanism) or in the mRNA sequence (in an mRNA-based mechanism), we explored whether this localization occurs in cells depleted of translating chloroplast ribosomes and LSU nascent chains through treatment with lincomycin (22,23). The rbcL FISH signal was not localized around the pyrenoid in cells treated with this drug (Fig.…”

Translation of chloroplast psbA mRNA is regulated by signals initiated by both photosystems II and I

“…However, the translation of psbD (encoding the D2 protein), rbcL (encoding the large subunit of ribulose bisphosphate carboxylase͞oxygenase), and psaA-psaB (encoding the 65-and 70-kDa chlorophyll a apoproteins of PS I) has been shown to be light-regulated as well (4,5,9). Both initiation and elongation steps of translation are probably controlled by light (1,5,(10)(11)(12)(13)(14)(15)(16).…”

“…The preservation of light-induced translation in isolated intact chloroplasts (10,17,18) suggests that the factors mediating light-signal perception and transduction are localized within this organelle. To date, only the light-capturing reactions of photosynthesis have been shown to act as photoreceptors within the chloroplast.…”

Translation of Chloroplast psbA mRNA Is Modulated in the Light by Counteracting Oxidizing and Reducing Activities

“…While in the mature chloroplast the demand for rapid response in gene expression, to accommodate an environment of fluctuating light intensities, probably selected translation as a major determinant of photoregulated gene expression (reviewed in references 11, 19, 39, and 53). Both initiation and elongation steps of translation have been implicated as light-regulated components of gene expression (5,17,27,29,56).…”