Comparison of the Molecular Weights of Proteins Synthesized by Isolated Chloroplasts with Those Which Appear during Greening in <i>Zea mays</i>

Cell-free protein synthesizing systems have been used to study the developmental changes in the synthesis of chloroplast proteins in the cotyledons of cucumber seedlings grown in the lght or in the dark. Escherichia coi and wheat germ in vitro protein synthesizing systems have been used to assay the changes in the levels of the mRNA's coding for ribulose 1,5-bisphosphate carboxylase (RuBPCase). The large subunit of cucumber RuBPCase has been identified among the translation products of the E. coli system. The wheat germ system translates the cucumber mRNA coding for the small subunit of RuBPCase to produce a 25,000 molecular weight precursor polypeptide. Plastids isolated from lght-grown cotyledons were used to study developmental changes in their capacity to synthesize protein. The data obtained indicate that in the light there is an initial 48-hour period of accumulation of the mRNA's coding for the large and small subunits of RuBPCase, coupled with an increase in the capacity of the isolated plastids to synthesize protein. This is followed by a decline. This decline is not reflected in the accumulation of RuBPCase in the cotyledons which remains constant over the period of study.The cotyledons of seeds which exhibit epigeal germination provide a useful system to study plastid development. As part of a program to study germination and development of cucumber (Cucumis sativus) seedlings (4) we became interested in the acquisition of photosynthetic competence by the cotyledons. During seedling development, the initial source of energy for the seedling is provided by the metabolism of stored.lipid (4). However, once the cotyledons emerge above the soil there is a decline in the activities of the enzymes that are involved in lipid metabolism. This decline coincides with increases in the levels of phototrophic indicators such as Chl and RuBPCase3 (4). Our aim was to study the control mechanisms involved in the synthesis of chloroplast proteins during the transiti6n of the cotyledon from growth on endogenous reserves to growth on newly fixed carbon.In this paper we describe the use ofcell-free protein synthesizing systems to study developmentally related changes in the levels of translatable mRNAs which code for RuBPCase within the cotyledons of cucumber seedlings. Previous

Section: Resultsmentioning

confidence: 49%

Synthesis of Chloroplast Proteins during Germination and Early Development of Cucumber

Walden¹,

Leaver²

1981

“…Specific and absolute requirements for light during development were found for some thylakoid membrane polypeptides in greening etioplasts (13). At least one of these polypeptides, the apoprotein of CPI, is known to be synthesized by chloroplast ribosomes (4).…”

Section: Tionmentioning

confidence: 99%

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

Fish¹,

Jagendorf²

1982

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-'

“…Plastids develop that resemble chloroplasts of immature green fruit, except that many ofthe plastoglobuli formed during differentiation of chloroplasts to chromoplasts are still present in the redifferentiated chloroplasts (23). Development grown seedlings contain the chloroplastic proteins RuBPCase (18,22,27) and CF, (7). Upon exposure to light, etioplasts rapidly become photosynthetically competent chloroplasts, accumulating Chl, thylakoids, and new polypeptides including LHCP and CPI (1,18,26,27).…”

mentioning

confidence: 99%

Accumulation of Chlorophyll, Chloroplastic Proteins, and Thylakoid Membranes during Reversion of Chromoplasts to Chloroplasts in Citrus sinensis Epicarp

Mayfield¹,

Huff²

1986

In vitro culture of pericarp segments from fruit of Citrus sinensis (L.) Osbeck cv Valencia was used to determine the temporal sequence in development of chloroplasts from chromoplasts during regreening of the epicarp. Regreening of chromoplasts closely resembled greening of etioplasts, except that regreening proceeded much more slowly. Chlorophyll, the light-harvesting chlorophyll a/b binding protein of photosystem II, the chlorophyll a binding protein of reaction center P-700 of photosystem I, thylakoid membranes, and adenosine triphosphate synthetase were all detected at very low levels in degreened epicarp. All of these increased in parallel during regreening of the epicarp. Ribulose Redifferentiation of chromoplasts to chloroplasts has been observed in regreening ofseveral plant organs (6,9,19) grown seedlings contain the chloroplastic proteins RuBPCase (18,22,27) and CF, (7). Upon exposure to light, etioplasts rapidly become photosynthetically competent chloroplasts, accumulating Chl, thylakoids, and new polypeptides including LHCP and CPI (1,18,26,27).Reversible interconversions between chloroplasts and chromoplasts can be conveniently studied by culturing pericarp segments from citrus fruit on agar media (11). Media with high sugar concentrations and lacking N promote differentiation of chloroplasts into chromoplasts, while media with low sugar concentrations or any of several sources of N promote redifferentiation of chromoplasts into chloroplasts.This paper presents results of experiments using cultured segments of pericarp from C. sinensis (L.) Osbeck cv Valencia to determine the temporal sequence of appearance of Chl, thylakoids, and certain chloroplastic proteins during redifferentiation of chloroplasts from chromoplasts. Chl, LHCP, CPI, and thylakoid membranes were present in very low abundance in chromoplasts ofC. sinensis. The reaccumulation ofthese components during development of chloroplasts from chromoplasts resembled chloroplast development from etioplasts, except for the time required. RuBPCase and CF, were present in chromoplasts of C. sinensis prior to regreening on agar media. RuBPCase levels decreased markedly for the first 10 d of culture, but reaccumulated in segments that regreened. Light was required for regreening and for the reaccumulation of RuBPCase after the initial loss. The absence of N from low sugar media reduced the accumulation of Chl, thylakoids, and Chl-binding proteins and completely prevented the reaccumulation of RuBPCase. High concentrations ofsucrose (150 mM) in the absence ofN inhibited the accumulation of LHCP and regreening, but this inhibition was largely overcome by the inclusion of KNO3 in the culture medium. MATERIALS AND METHODSPericarp Culture. Segments of pericarp 1 cm in diameter were prepared from fruit of Citrus sinensis (L.) Osbeck cv Valencia as previously described (1 1). The segments were kept on agar media described by Murashigi and Tucker (17), except that 60 mm KN03 was the sole source of N, the sucrose concentration was reduced to 1...