Synthesis of a chloroplast membrane polypeptide on thylakoid-bound ribosomes during the cell cycle of Chlamydomonas reinhardii 137+

Herrin, David L.; Michaels, Allan; Hickey, Eileen

doi:10.1016/0005-2787(81)90003-4

Cited by 42 publications

(21 citation statements)

References 25 publications

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“…We have to assume that the very small amounts actually needed came in as contaminants, especially with the S-30 which was only briefly desalted. The optima at 0.2 mm for GTP and 1.0 mm for ATP are very close to concentrations used in previous work (3,15,21 The absence of any effect of peptide chain initiation inhibitors (Table II) agrees with two earlier reports of the lack of inhibition of ATA (3,13). Bolli et aL (5) reported 35% inhibition ofprotein synthesis by washed thylakoids from Chlamydomonas due to 1 00,M ATA; but this concentration is likely to inhibit elongation as well (26).…”

Section: Resultssupporting

confidence: 89%

“…1) was remarkably sharp, and corresponds well with the 80 mm needed by broken spinach chloroplasts (6). Herrin et al (13) used 70 mm K+ together with 40 mm NH4' for translation by thylakoid-bound ribosomes of Chlamydomonas, with the total close to the 100 mm optimum we find, but did not calculate absolute rates of leucine incorporation. Alscher et al (3) and Margulies and Michaels (20) used 120 and 50 mM KCI, respectively, far enough away from the optimum, to account, in part, for the lower rates of incorporation they reported.…”

Section: Resultssupporting

confidence: 74%

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Optimal Conditions for Translation by Thylakoid-Bound Polysomes from Pea Chloroplasts

Bhaya

Jagendorf²

1984

Plant Physiol.

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ABSTRACIPolysomes bound to washed thylakoids from pea Pisum sativum cv Progress No. 9 chloroplasts are capable of protein synthesis when supplemented with amino acids, ATP and a regenerating system, GTP, and soluble factors required for translation. The extent of protein synthesis in previous reports, however, was quite low when compared to in organeilo transltion. By systematic testing of parameters in the isolation of thylakoids and reaction mixture components we have been able to establish more optimal conditions. Incorporation of 2 to 10 nanomoles of leucine per milligram chlorophyll in a 20-minute reaction period is now possible, representing a 10-to 60-fold increase over amounts previously reported. Autoradiographs of solubilized, electrophoresed membranes show about 30 discrete labeled polypeptides which remain associated with the thylakoid membranes.There are two populations of ribosomes within chloroplastthose free in the stroma, and those bound to the thylakoid membranes (20,27). Further, it is known that a large fraction of the bound ribosomes are in the form of polysomes attached at least in part by nascent peptide chains (3,20,21,27). These attached polysomes can continue protein synthesis when supplied with ATP, GTP, amino acids, and soluble factors (enzymes and tRNAs) for translation, although at quite low rates (3, 21) compared to those found in organello (12,22). Before investigating the nature of polypeptides synthesized by attached polysomes, we felt it important to optimize the conditions for translation, since a distorted picture of the extent and relative abundance of specific polypeptides may result from nonoptimal reaction rates (17

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Section: Resultssupporting

confidence: 89%

Section: Resultssupporting

confidence: 74%

Optimal Conditions for Translation by Thylakoid-Bound Polysomes from Pea Chloroplasts

Bhaya

Jagendorf²

1984

Plant Physiol.

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“…Western blot analyses (data not shown) identify peak 2 as the D2 protein (7) of P511. We speculate that photoinhibition rapidly modifies the D2 protein, and that it is slowly excised and replaced with newly synthesized D2 protein (11,16) in vivo during slow (t112 '-3 h), light-dependent recovery from photoinhibition. Apparently the 60 mmn period of photoinhibition of [35S]-prelabeled NH2OH extracted leaves in the experiment of Figure 6 was too short to detect excision of the D2 protein.…”

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confidence: 94%

Studies on the Photoactivation of the Water-Oxidizing Enzyme

Callahan¹,

Becker²,

Cheniae³

1986

Plant Physiol.

134

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Inactivation of the water splitting enzyme complex in leaves or isolated chloroplasts results in increased susceptibility of photosystem II (PSII) to damage by light. Photoinhibition under this condition occurs in very weak light. The site of damage is exclusive of the water splitting complex yet still on the oxidizing side of PSII, as the QB locus is unaffected while photoreduction of silicomolybdate is inhibited. The kinetics of loss in PSII activity are more complex than apparent first-order, and the quantum efficiency is low. We observe no evidence of deletion from thylakoid membranes of any PSII polypeptide as a consequence of photoinhibition, although recovery from the photoinhibition is dependent upon both light and 70S protein synthesis. Enhanced synthesis of two proteins occurs during recovery, only one of which (D2) appears to be causally related to the recovery. We present a model which describes the relationship of weak light photoinhibition and its recovery to photoactivation of the S-state water oxidizing complex.Irradiation of oxygenic photosynthetic organisms or their isolated chloroplasts with intense visible light inhibits photosynthesis by poorly understood processes at specific loci in the electron transport sequence and reaction centers (see Refs. 33 and 38 for recent reviews). Such photoinhibitions of photosynthesis proceed via low quantum yield processes following light absorption by Chl (18)

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“…In contrast, all radioactive products synthesized by thylakoid-bound ribosomes remained with the membrane even after puromycin treatment, and hence, were thought to be hydrophobic polypeptides. Subsequently, several workers (Alscher et al 1978, Herrin et al 1981, Herrin and Michaels 1985, Margulies 1983, Bhaya and Jagendorf 1984 confirmed that the "function of thylakoid-bound ribosomes is to synthesize thylakoid membrane proteins" (Herrin et al 1981). The finding that the extrinsic a-and /?-subunits of CF, are synthesized on both free and membrane-bound polysomes of pea chloroplasts (Bhaya and Jagendorf 1985) supported the idea that the hydrophobicity of the proteins determines their site of synthesis.…”

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confidence: 95%

Quantification of mRNA in Chloroplasts of <italic>Chlamydomonas reinhardii</italic>: Equal Distribution of mRNA for a Soluble and a Membrane Polypeptide in Stroma and Thylakoids

Breidenbach

Jenni

Leu

et al. 1988

Plant and Cell Physiology

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The relative contents of the mRNAs were analyzed for the 32 kDa herbicide-binding protein and for the large subunit of ribulose-l,5-bisphosphate carboxylase in the membrane fraction and in the soluble fraction of chloroplasts from Chlamydomonas reinhardii. The presence of mRNA for the two proteins in both subchloroplast fractions was demonstrated by in vitro translation of isolated RNA in the reticulocyte lysate. The relative amounts of the two mRNAs were measured by hybridizations with cloned chloroplast DNA probes at two stages of the cell cycle. Both mRNAs were distributed in the same ratio between membrane and soluble fractions, about 75% of both mRNAs being in the membrane and 25% in the soluble fraction. Therefore, in chloroplasts the accumulation of mRNAs on thylakoid membranes does not reflect the final localization of soluble and membrane proteins.

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Synthesis of a chloroplast membrane polypeptide on thylakoid-bound ribosomes during the cell cycle of Chlamydomonas reinhardii 137+

Cited by 42 publications

References 25 publications

Optimal Conditions for Translation by Thylakoid-Bound Polysomes from Pea Chloroplasts

Optimal Conditions for Translation by Thylakoid-Bound Polysomes from Pea Chloroplasts

Studies on the Photoactivation of the Water-Oxidizing Enzyme

Quantification of mRNA in Chloroplasts of <italic>Chlamydomonas reinhardii</italic>: Equal Distribution of mRNA for a Soluble and a Membrane Polypeptide in Stroma and Thylakoids

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