scholarly journals Substrate- and species-specific processing enzymes for chloroplast precursor proteins

1994 ◽  
Vol 300 (3) ◽  
pp. 787-792 ◽  
Author(s):  
Q Su ◽  
A Boschetti
Keyword(s):  
Higher Plants ◽  
Gel Filtration ◽  
Molecular Size ◽  
Small Subunit ◽  
Bisphosphate Carboxylase ◽  
Chlamydomonas Reinhardii ◽  
Processing Enzymes ◽  
Precursor Proteins ◽  
Species Specific ◽  
Correct Size

Using different precursors of chloroplast proteins and stromal extracts from both Chlamydomonas reinhardii and pea chloroplasts, we analysed the specificity of stroma-localized processing peptidases. By gel filtration of a stromal extract from isolated Chlamydomonas chloroplasts, fractions could be separated containing enzymic activities for processing the precursors of the small subunit of ribulose-1,5-bisphosphate carboxylase (pSS) and of the protein OEE1 from the photosynthetic water-splitting complex (pOEE1). The enzymes differed not only in molecular size, but also in their sensitivity to inhibitors and in their pH optima. Obviously, in the stroma of Chlamydomonas chloroplasts different peptidases exist for processing of pSS and pOEE1, the latter being converted into an intermediate-sized form, iOEE1, which was found to be further processed to mature OEE1 by a thylakoid-associated protease. To study the species-specificity of the stromal peptidases, stromal extracts from Chlamydomonas and pea chloroplasts were incubated with pSS from either of these organisms. In the heterologous combinations, the precursors were partly hydrolysed, but not to the correct size. In importation assays, pSS from pea (but also the precursor of the ribosomal protein L12 from spinach) could not enter into chloroplasts from Chlamydomonas. In contrast, the algal pSS was imported into chloroplasts from pea, although it was not processed to mature SS. Our results indicate that the importation machinery and the pSS-processing enzymes in higher plants and green algae have different specificities and that in Chlamydomonas several stromal peptidases for different precursor proteins exist.

scholarly journals Two components of the chloroplast protein import apparatus, IAP86 and IAP75, interact with the transit sequence during the recognition and translocation of precursor proteins at the outer envelope.

1996 ◽  
Vol 134 (2) ◽  
pp. 315-327 ◽  
Author(s):  
Y Ma ◽  
A Kouranov ◽  
S E LaSala ◽  
D J Schnell
Keyword(s):  
Outer Membrane ◽  
Protein Import ◽  
Small Subunit ◽  
Chloroplast Envelope ◽  
Stable Complex ◽  
Nucleoside Triphosphate ◽  
Cross Linking ◽  
Outer Envelope ◽  
Bisphosphate Carboxylase ◽  
Precursor Proteins

The interactions of precursor proteins with components of the chloroplast envelope were investigated during the early stages of protein import using a chemical cross-linking strategy. In the absence of energy, two components of the outer envelope import machinery, IAP86 and IAP75, cross-linked to the transit sequence of the precursor to the small subunit of ribulose-1, 5-bisphosphate carboxylase (pS) in a precursor binding assay. In the presence of concentrations of ATP or GTP that support maximal precursor binding to the envelope, cross-linking to the transit sequence occurred predominantly with IAP75 and a previously unidentified 21-kD polypeptide of the inner membrane, indicating that the transit sequence had inserted across the outer membrane. Cross-linking of envelope components to sequences in the mature portion of a second precursor, preferredoxin, was detected in the presence of ATP or GTP, suggesting that sequences distant from the transit sequence were brought into the vicinity of the outer membrane under these conditions. IAP75 and a third import component, IAP34, were coimmunoprecipitated with IAP86 antibodies from solubilized envelope membranes, indicating that these three proteins form a stable complex in the outer membrane. On the basis of these observations, we propose that IAP86 and IAP75 act as components of a multisubunit complex to mediate energy-independent recognition of the transit sequence and subsequent nucleoside triphosphate-induced insertion of the transit sequence across the outer membrane.

Photoregulation of the biosynthesis of ribulose bisphosphate carboxylase

Development ◽  
1984 ◽  
Vol 83 (Supplement) ◽  
pp. 163-178
Author(s):  
R. John Ellis ◽  
Thomas F. Gallagher ◽  
Gareth I. Jenkins ◽  
C. Ruth Lennox
Keyword(s):  
Chloroplast Development ◽  
Higher Plants ◽  
Large Subunit ◽  
Nuclear Genome ◽  
Small Subunit ◽  
Ribulose Bisphosphate Carboxylase ◽  
Ribulose Bisphosphate ◽  
Bisphosphate Carboxylase ◽  
Subunit Mrna ◽  
Parallel Increase

Chloroplast development in higher plants is light dependent, and is accompanied by the synthesis of chlorophyll and the accumulation of many chloroplast polypeptides. There is a 100-fold greater content of the photosynthetic enzyme, ribulose-1,5-bisphosphate carboxylase-oxygenase, in light-grown seedlings of Pisum sativum than in dark-grown seedlings. Following the illumination of dark-grown seedlings, there is a parallel increase in the content of both the mRNA and the polypeptide of the small subunit of the carboxylase; this subunit is a product of the nuclear genome. The increases in the mRNA and the polypeptide of the large subunit, which is a product of the chloroplast genome, show less synchronicity. Studies with isolated leaf nuclei show that the increase in small subunit mRNA is mediated primarily at the level of transcription. Three distinct effects of light on transcription of small subunit genes have been found; a rapid (∼1 h) burst, followed by a decline, when etiolated plants are first exposed to light; a slow (∼36h) development of the competence to transcribe rapidly after the initial burst; rapid (∼20 min) switches in both directions when fully greened plants are exposed to light—dark transitions.

scholarly journals Condensation of Rubisco into a proto-pyrenoid in higher plant chloroplasts

2020 ◽  
Author(s):  
Nicky Atkinson ◽  
Yuwei Mao ◽  
Kher Xing Chan ◽  
Alistair J. McCormick
Keyword(s):  
Higher Plants ◽  
Initial Step ◽  
Small Subunit ◽  
Operating Efficiency ◽  
Higher Plant ◽  
Bisphosphate Carboxylase ◽  
Spontaneous Condensation ◽  
Concentrating Mechanism ◽  
Chloroplast Stroma ◽  
Almost All

SummaryPhotosynthetic CO2 fixation in plants is limited by the inefficiency of the CO2-assimilating enzyme Rubisco (D-ribulose-1,5-bisphosphate carboxylase/ oxygenase)1–3. In plants possessing the C3 pathway, which includes most major staple crops, Rubisco is typically evenly distributed throughout the chloroplast stroma. However, in almost all eukaryotic algae Rubisco aggregates within a microcompartment known as the pyrenoid, in association with a CO2-concentrating mechanism that improves photosynthetic operating efficiency under conditions of low inorganic carbon4. Recent work has shown that the pyrenoid matrix is a phase-separated, liquid-like condensate5. In the alga Chlamydomonas reinhardtii, condensation is mediated by two components: Rubisco and the linker protein EPYC1 (Essential Pyrenoid Component 1)6,7. Here we show that expression of mature EPYC1 and a plant-algal hybrid Rubisco leads to spontaneous condensation of Rubisco into a single phase-separated compartment in Arabidopsis chloroplasts, with liquid-like properties similar to a pyrenoid matrix. The condensate displaces the thylakoid membranes and is enriched in hybrid Rubisco containing the algal Rubisco small subunit required for phase separation. Promisingly, photosynthetic CO2 fixation and growth is not impaired in stable transformants compared to azygous segregants. These observations represent a significant initial step towards enhancing photosynthesis in higher plants by introducing an algal CO2-concentrating mechanism, which is predicted to significantly increase the efficiency of photosynthetic CO2 uptake8,9.

The Rubisco small subunit gene as a paradigm for studies on differential gene expression during plant development

1986 ◽  
Vol 313 (1162) ◽  
pp. 409-417 ◽  
Keyword(s):  
Transcriptional Control ◽  
Higher Plants ◽  
Mrna Level ◽  
Small Subunit ◽  
Upstream Sequence ◽  
Specific Expression ◽  
Bisphosphate Carboxylase ◽  
Cis Acting ◽  
Transgenic Petunia ◽  
Light Inducibility

The ribulose 1,5-bisphosphate carboxylase small subunit (rbcS) in higher plants is encoded by a small multigene family. Members of the gene family contain 1-3 introns. The rbcS mRNA is differentially distributed in various plant organs. It is most abundant in leaves, less so in stems and other photosynthetic organs, and almost undetectable in roots. In leaves, the rbcS mRNA level is greatly increased by light through transcriptional control of the genes. Ti-mediated gene transfer experiments have demonstrated that the pea rbcS-E9 gene retains light-regulated expression in transformed petunia calluses and in leaves of transgenic petunia and tobacco plants. A 33-base pair sequence around the TATA box region has been shown to be involved in the light-inducibility of the rbcS-E9 gene in transformed calluses. In transgenic petunia plants, the experiments thus far have shown that 352 base pairs of 5' upstream sequence is sufficient for light-inducibility, as well as for leaf-specific expression. Further experiments in progress will help to identify and characterize other cis-acting elements involved in the differential expression of the rbcS genes.

Cis -acting elements for selective expression of two photosynthetic genes in transgenic plants

1986 ◽  
Vol 314 (1166) ◽  
pp. 493-500 ◽  
Keyword(s):  
Higher Plants ◽  
Small Subunit ◽  
Upstream Region ◽  
Organ Specificity ◽  
Base Pairs ◽  
Bisphosphate Carboxylase ◽  
Cis Acting ◽  
Conserved Sequence ◽  
Photosynthetic Genes

The most abundant mRNAs in leaves of higher plants encode the small subunit (rbcS) of ribulose 1,5-bisphosphate carboxylase and the chlorophyll a/b -binding (Cab) protein. Nuclear genes for these mRNAs are expressed in an organ-specific manner and their expression is induced by light acting through phytochrome. We have used DNA sequence manipulation in vitro coupled with transgenic plant expression systems to define sequence determinants for regulated expression of these two photosynthetic genes. At least two cis -acting elements for the rbcS genes have been identified. A conserved sequence from the transcription start site to the 5' boundary of the TATA box is involved in light-inducible transcription. In addition, an upstream enhancer-like element (240-280 base pairs) confers phytochrome responsiveness and organ specificity on constitutive promoters. An enhancer-like element ( ca . 260 base pairs) in the upstream region of the wheat Cab -1 gene also shows similar functions.

Specific binding of acrosome-reaction-inducing substance to the head of starfish spermatozoa

Zygote ◽  
1993 ◽  
Vol 1 (2) ◽  
pp. 121-127 ◽  
Author(s):  
Akira Ushiyama ◽  
Takeo Araki ◽  
Kazuyoshi Chiba ◽  
Motonori Hoshi
Keyword(s):  
Acrosome Reaction ◽  
Specific Binding ◽  
Molecular Size ◽  
Polystyrene Latex ◽  
Anterior Region ◽  
Head Region ◽  
Signal Molecule ◽  
Jelly Coat ◽  
Plot Analysis ◽  
Species Specific

In the starfish, spermatozoa undergo the acrosome reaction upon encountering the jelly coat of eggs. A highly sulphated glycoprotein in the jelly coat is called acrosome-reaction-inducing substance (ARIS) because it is the key signal molecule to trigger the acrosome reaction. The activity of ARIS is mainly attributed to its sulphate and saccharide residues. The extremely large molecular size and speciesspecific action of ARIS suggest the presence of a specific ARIS receptor on the sperm surface, but no experimental evidence for the receptor has been presented. We therefore measured specific binding of ARIS and its pronase digest (P-ARIS), which retains the full activity of ARIS, to homologous spermatozoa by using fluorescien-isothiocyanate-labelled ARIS and125 I-labelled P-ARIS, respectively. The spermatozoa had the ability to bind ARIS, as well as P-ARIS, specifically. The binding was species-specific, and mostly localised to the head region of spermatozoa. Scatchard plot analysis indicated the presence of one class of ARIS receptor on the surface of acrosome-intact speramatozoa. Furthermore, the specific binding of P-ARIS to the anterior region of sperm heads was microscopically confirmed by using P-ARIS conjugated to polystyrene latex beads with intense fluorescence. It is concluded that starfish spermatozoa have a specific receptor for ARIS on the surface of the anterior region of heads.

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