14-3-3 proteins control proteolysis of nitrate reductase in spinach leaves.
(57/9955)
To test a possible role of 14-3-3 proteins in the degradation of nitrate reductase (NR) in leaves, we monitored 14-3-3s bound to NR in leaf extracts. The amount of 14-3-3s that coimmunoprecipitated with serine 543 phospho-NR (p-NR) increased upon a light/dark transition. This was accompanied by a similar increase in the protein turnover rate of NR in leaves. Purified NR was degraded in extracts from darkened but not from illuminated leaves. Removal of 14-3-3s from such extracts prevented NR degradation. We conclude that the availability of 14-3-3s for p-NR regulates the stability of NR. (+info)
Subpopulations of chloroplast ribosomes change during photoregulated development of Zea mays leaves: ribosomal proteins L2, L21, and L29.
(58/9955)
Seedlings grown in darkness, i.e., etiolated seedlings, lack chlorophyll and most other components of the photosynthetic apparatus. On illumination, the plastids become photosynthetically competent through the production of chlorophylls and proteins encoded by certain chloroplast and nuclear genes. There are two types of photosynthetic cells in leaves of the C4 plant maize: bundle sheath cells (BSC) and adjacent mesophyll cells (MC). Some proteins of the maize photosynthetic machinery are solely or preferentially localized in MC and others in BSC. A particular gene may be photoregulated up in one cell type and down in the other. Transcripts of the nuclear gene rpl29, encoding the chloroplast ribosomal protein L29, increase in abundance about 17-fold during light-induced maturation of plastids. There is about 1.5 times more L29 protein in ribosomes of greening leaves than in ribosomes of unilluminated leaves; the L29 contents of MC and BSC are about the same. However, L21 is present about equally in plastid ribosomes of unilluminated and illuminated seedlings. In contrast to both L29 and L21, the fraction of the ribosome population containing L2 is about the same in MC and BSC of etiolated leaves but, on illumination, the proportion of the ribosome population with L2 increases in BSC but not in MC. The existence of different subpopulations of plastid ribosomes-e.g., those with and without L21 and/or L29 during development-evokes interesting, but as yet unanswered, questions about the roles of different types of ribosomes in differentiation. (+info)
Changes in the shapes of leaves and flowers upon overexpression of cytochrome P450 in Arabidopsis.
(59/9955)
In Arabidopsis, the two-dimensional expansion of leaves is regulated via the polarized elongation of cells. The ROTUNDIFOLIA3 (ROT3) protein, a member of the family of cytochromes P450, is involved in this process and regulates leaf length. Transgenic plants that overexpressed a wild-type ROT3 gene had longer leaves than parent plants, without any changes in leaf width. The shapes of floral organs were also altered, but elongation of the stem, roots, and hypocotyls was unaffected. To our knowledge, no similar specific regulation of leaf length has been reported previously. Transgenic plants overexpressing the rot3-2 gene had enlarged leaf blades but leaf petioles of normal length. Morphological alterations in such transgenic plants were associated with changes in shape of leaf cells. The ROT3 gene seems to play an important role in the polar elongation of leafy organs and should be a useful tool for the biodesign of plant organs. (+info)
Senescence-associated gene expression during ozone-induced leaf senescence in Arabidopsis.
(60/9955)
The expression patterns of senescence-related genes were determined during ozone (O(3)) exposure in Arabidopsis. Rosettes were treated with 0.15 microL L(-1) O(3) for 6 h d(-1) for 14 d. O(3)-treated leaves began to yellow after 10 d of exposure, whereas yellowing was not apparent in control leaves until d 14. Transcript levels for eight of 12 senescence related genes characterized showed induction by O(3). SAG13 (senescence-associated gene), SAG21, ERD1 (early responsive to dehydration), and BCB (blue copper-binding protein) were induced within 2 to 4 d of O(3) treatment; SAG18, SAG20, and ACS6 (ACC synthase) were induced within 4 to 6 d; and CCH (copper chaperone) was induced within 6 to 8 d. In contrast, levels of photosynthetic gene transcripts, rbcS (small subunit of Rubisco) and cab (chlorophyll a/b-binding protein), declined after 6 d. Other markers of natural senescence, SAG12, SAG19, MT1 (metallothionein), and Atgsr2 (glutamine synthetase), did not show enhanced transcript accumulation. When SAG12 promoter-GUS (beta-glucuronidase) and SAG13 promoter-GUS transgenic plants were treated with O(3), GUS activity was induced in SAG13-GUS plants after 2 d but was not detected in SAG12-GUS plants. SAG13 promoter-driven GUS activity was located throughout O(3)-treated leaves, whereas control leaves generally showed activity along the margins. The acceleration of leaf senescence induced by O(3) is a regulated event involving many genes associated with natural senescence. (+info)
Phosphoenolpyruvate carboxykinase in the C(4) monocot Urochloa panicoides is encoded by four differentially expressed genes.
(61/9955)
Previous screening of a cDNA library of leaf poly(A(+)) RNA from Urochloa panicoides, a phosphoenolpyruvate carboxykinase (PCK)-type C(4) monocot, led to the characterization of cDNAs encoding the U. panicoides PCK subunit PCK1. A second PCK sequence, designated PCK2, has now been found by rescreening the library. The deduced PCK2 polypeptide is 626 residues in length, has a predicted molecular mass of 68,686 D, and is 96% identical to the deduced PCK1 sequence. Isolation and characterization of genomic DNA fragments revealed that the PCK1 and PCK2 genes are each closely linked to another PCK gene. These additional genes have been designated PCK3 and PCK4, respectively. In each case, the second gene is located upstream and in the same transcriptional orientation as the gene characterized through cDNA analysis. A reverse transcription-polymerase chain reaction assay was used to demonstrate that PCK1 and PCK2 transcripts predominate in leaves, whereas PCK3 and PCK4 transcripts predominate in roots. Moreover, accumulation of PCK1 and PCK2 transcripts is light dependent. Direct N-terminal sequencing of PCK polypeptides purified from leaves demonstrated that PCK2 is produced. These results strongly suggest that PCK1 and PCK2 are involved in the photosynthetic CO(2)-concentrating mechanism active in U. panicoides. (+info)
S-methylmethionine plays a major role in phloem sulfur transport and is synthesized by a novel type of methyltransferase.
(62/9955)
All flowering plants produce S-methylmethionine (SMM) from Met and have a separate mechanism to convert SMM back to Met. The functions of SMM and the reasons for its interconversion with Met are not known. In this study, by using the aphid stylet collection method together with mass spectral and radiolabeling analyses, we established that l-SMM is a major constituent of the phloem sap moving to wheat ears. The SMM level in the phloem ( approximately 2% of free amino acids) was 1.5-fold that of glutathione, indicating that SMM could contribute approximately half the sulfur needed for grain protein synthesis. Similarly, l-SMM was a prominently labeled product in phloem exudates obtained by EDTA treatment of detached leaves from plants of the Poaceae, Fabaceae, Asteraceae, Brassicaceae, and Cucurbitaceae that were given l-(35)S-Met. cDNA clones for the enzyme that catalyzes SMM synthesis (S-adenosylMet:Met S-methyltransferase; EC 2.1.1.12) were isolated from Wollastonia biflora, maize, and Arabidopsis. The deduced amino acid sequences revealed the expected methyltransferase domain ( approximately 300 residues at the N terminus), plus an 800-residue C-terminal region sharing significant similarity with aminotransferases and other pyridoxal 5'-phosphate-dependent enzymes. These results indicate that SMM has a previously unrecognized but often major role in sulfur transport in flowering plants and that evolution of SMM synthesis in this group involved a gene fusion event. The resulting bipartite enzyme is unlike any other known methyltransferase. (+info)
Subunit C of the vacuolar H+-ATPase of Hordeum vulgare.
(63/9955)
The molecular cloning of the first subunit C of the plant vacuolar H+-ATPase is reported. Tonoplast vesicles were purified from barley leaves by sucrose gradient centrifugation, and the tonoplast polypeptides were separated by two-dimensional (2-D) gel electrophoresis. Using an anti-ATPase holoenzyme antibody, a polypeptide was recognized in the molecular mass range of 40 kDa with an isoelectric point of about 6.0, and tentatively identified as subunit C. The polypeptide spot was excised from about 50 2-D gels and subjected to endo Lys C proteolysis. Two proteolytic peptides were sequenced and the amino acid sequences were used to design degenerated oligonucleotides, followed by PCR amplification with cDNA template and screening of a cDNA library synthesized from Hordeum vulgare poly A mRNA of epidermis strips. The full length clone of 1.5 kbp contains an open reading frame of 1062 bp encoding a polypeptide of 354 amino acids with a molecular mass of 39,982 Da and an isoelectric point of 6.04. Amino acid identity with sequences of SUC from animals and fungi is in the range of 36.7 to 38.5%. Expression of the cloned gene was demonstrated by Northern blotting and RT-PCR. (+info)
Isolation and characterisation of the cDNA encoding a glycosylated accessory protein of pea chloroplast DNA polymerase.
(64/9955)
The cDNA encoding p43, a DNA binding protein from pea chloroplasts (ct) that binds to cognate DNA polymerase and stimulates the polymerase activity, has been cloned and characterised. The characteristic sequence motifs of hydroxyproline-rich glyco-proteins (HRGP) are present in the cDNA corres-ponding to the N-terminal domain of the mature p43. The protein was found to be highly O-arabinosylated. Chemically deglycosylated p43 (i.e. p29) retains its binding to both DNA and pea ct-DNA polymerase but fails to stimulate the DNA polymerase activity. The mature p43 is synthesised as a pre-p43 protein containing a 59 amino acid long transit peptide which undergoes stromal cleavage as evidenced from the post-translational in vitro import of the precursor protein into the isolated intact pea chloroplasts. Surprisingly, p43 is found only in pea chloroplasts. The unique features present in the cloned cDNA indicate that p43 is a novel member of the HRGP family of proteins. Besides p43, no other DNA-polymerase accessory protein with O-glycosylation has been reported yet. (+info)