Rough Thylakoids: Polysomes Attached to Chloroplast Membranes

Falk, H.

doi:10.1083/jcb.42.2.582

Cited by 51 publications

(10 citation statements)

References 38 publications

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“…, 1963; Eilam et al. , 1971; Gunning and Steer, 1975), and a significant proportion of the ribosomes (variously estimated at 20–50%) is associated with the thylakoid membranes (Falk, 1969; Chua et al. , 1973, 1976; Yamamoto et al.…”

Section: Discussionmentioning

confidence: 99%

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Exclusion of plastid nucleoids and ribosomes from stromules in tobacco and Arabidopsis

et al. 2011

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SUMMARYStromules are stroma-filled tubules that extend from the surface of plastids and allow the transfer of proteins as large as 550 kDa between interconnected plastids. The aim of the present study was to determine if plastid DNA or plastid ribosomes are able to enter stromules, potentially permitting the transfer of genetic information between plastids. Plastid DNA and ribosomes were marked with green fluorescent protein (GFP) fusions to LacI, the lac repressor, which binds to lacO-related sequences in plastid DNA, and to plastid ribosomal proteins Rpl1 and Rps2, respectively. Fluorescence from GFP-LacI co-localised with plastid DNA in nucleoids in all tissues of transgenic tobacco (Nicotiana tabacum L.) examined and there was no indication of its presence in stromules, not even in hypocotyl epidermal cells, which contain abundant stromules. Fluorescence from Rpl1-GFP and Rps2-GFP was also observed in a punctate pattern in chloroplasts of tobacco and Arabidopsis [Arabidopsis thaliana (L.) Heynh.], and fluorescent stromules were not detected. Rpl1-GFP was shown to assemble into ribosomes and was co-localised with plastid DNA. In contrast, in hypocotyl epidermal cells of dark-grown Arabidopsis seedlings, fluorescence from Rpl1-GFP was more evenly distributed in plastids and was observed in stromules on a total of only four plastids (<0.02% of the plastids observed). These observations indicate that plastid DNA and plastid ribosomes do not routinely move into stromules in tobacco and Arabidopsis, and suggest that transfer of genetic information by this route is likely to be a very rare event, if it occurs at all.

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Section: Discussionmentioning

confidence: 99%

“…The confocal images were unable to distinguish thylakoid‐bound ribosomes, which have been observed by electron microscopy (Falk, 1969; Chua et al. , 1973, 1976; Yamamoto et al.…”

Section: Discussionmentioning

confidence: 99%

Exclusion of plastid nucleoids and ribosomes from stromules in tobacco and Arabidopsis

et al. 2011

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“…Some of the chloroplast 70S ribosomes are seen in electron micrographs to be associated with the thylakoid membranes (5,9,19,28). At least some of these ribosomes must be associated with mRNA, since washed thylakoids are capable of synthesizing proteins when supplemented with soluble factors (1,2,8,20).…”

Section: Tionmentioning

confidence: 99%

Light-Induced Increase in the Number and Activity of Ribosomes Bound to Pea Chloroplast Thylakoids in Vivo

Fish¹,

Jagendorf²

1982

Plant Physiol.

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Some of the chloroplast 70S ribosomes are seen in electron micrographs to be associated with the thylakoid membranes (5, 9, 19, 28). At least some of these ribosomes must be associated with mRNA, since washed thylakoids are capable of synthesizing proteins when supplemented with soluble factors (1,2,8,20). Retention in the thylakoids of the newly synthesized peptides provided evidence that the proteins synthesized were discharged into the membranes (2,20). From this, it seemed likely that the ribosomes active in vitro were tightly bound to the thylakoids by a nascent peptide chain. The presence of this type of linkage was demonstrated more directly for Chlamydomonas (5) and for pea (28), using puromycin-induced release of the ribosomes in the presence of high salt concentrations (0.5 to 1.0 M KCI) as a criterion.The nascent peptide linkages between ribosomes and the thy-'

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“…membranes and are likely to be specifically involved with the synthesis of membrane proteins. The association has been observed in electron microscope studies of Phaseolus leaves (8) and of Chlamydomonas (5). Even after isolated thylakoid membranes are washed repeatedly, ribosomes remain bound (1,5,9,15,16,30) and proteins synthesized by bound polysomes remain as part of the membrane system (1,4,7,19).…”

mentioning

confidence: 96%

Bound Ribosomes of Pea Chloroplast Thylakoid Membranes: Location and Release in Vitro by High Salt, Puromycin, and RNase

Yamamoto¹,

Burke²,

Autz³

et al. 1981

Plant Physiol.

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The mode of attachment of 70S ribosomes to thylakoid membranes from pea leaves was studied by determining the proportion of the bound RNA which was released by various incubation conditions. The results supported a model in which several classes of bound ribosomes could be distinguished:(a) very tightly bound, not released by any conditions yet tested (20% of the total); (b) monomeric ribosomes attached by electrostatic interaction with the membranes (30 to 40% of the total) and released by high salt; and (c) polysomes, with some of the ribosomes attached by a combination of electrostatic interactions and insertion of the nascent polypeptide chain into the membrane. These required a combination of puromycin and high salt for release. Other ("hanging") ribosomes of the polysomes were inferred to be attached through mRNA but not actually attached to the membranes directly; they could be released by RNase under low salt conditions, as weDl as by puromycin plus high salt.To obtain these results, chloroplasts had to be prepared in media containing 0.2 molar Tris at pH 8.5. Using Tricine buffers at pH 7.5 yielded thylakoid membranes whose ribosomes were removed almost completely by high salt alone; these showed no response to puromycin. However, pH 7.5 had to be used in all cases for ribosome dissociation in high salt media, as the ribosome structure appeared to be degraded by high salt at pH 8.5, and release then occurred without the need for puromycin.The kinetics of ribosome release by high salt showed a rapid initial phase with a half-life of 20 seconds. The extent of release by high salt was very dependent on the temperature of the incubation. Plotting the data according to the Arrhenius interpretation shows a significant break at about 15 C, with apparent activation energy of 20 kilocalories per mole below that temperature and 5 kilocalories per mole above that temperature.This result suggests that membrane fluidity might be an important factor permitting release of ribosomes under high salt conditions. Electron microscope pictures of the washed thylakoids showed polysomes closely associated with the outer membranes of grana stacks, and with the stroma lamellae. Following digitonin treatment of the membranes and centrifugation, fractions enriched in Photosystem I and presumed stroma lamellae were also enriched in bound RNA.Whereas many of the chloroplast 70S ribosomes are free in the stroma, others appear to be closely associated with the thylakoid ' Supported in part by Grant PCM-77-18839 membranes and are likely to be specifically involved with the synthesis of membrane proteins. The association has been observed in electron microscope studies of Phaseolus leaves (8) and of Chlamydomonas (5). Even after isolated thylakoid membranes are washed repeatedly, ribosomes remain bound (1,5,9,15,16,30) and proteins synthesized by bound polysomes remain as part of the membrane system (1,4,7,19). Estimates of the bound ribosomes as a per cent of the total number of plastid 70S ribosomes, measured in vitro, rang...

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Rough Thylakoids: Polysomes Attached to Chloroplast Membranes

Cited by 51 publications

References 38 publications

Exclusion of plastid nucleoids and ribosomes from stromules in tobacco and Arabidopsis

Exclusion of plastid nucleoids and ribosomes from stromules in tobacco and Arabidopsis

Light-Induced Increase in the Number and Activity of Ribosomes Bound to Pea Chloroplast Thylakoids in Vivo

Bound Ribosomes of Pea Chloroplast Thylakoid Membranes: Location and Release in Vitro by High Salt, Puromycin, and RNase

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