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M. Di Paola Lipucci D. Fossi F. Tognoni V. Nuti Ronchi 《Plant Cell, Tissue and Organ Culture》1987,10(1):3-10
Juvenile leaves of Cupressus arizonica Green (3–5 mm in length) from eight week old seedlings, were cultured on liquid medium supplemented with isopentenyladenine (2 mgl-1). Buds formed from the explants after three weeks of culture, but further growth occurred only after transfer to half-strength medium without plant growth regulators. Histological analysis at different times of culture, showed an early mitotic activity within transfusion tissue, followed by dedifferentiation of epidermal and mesophyll parenchyma cells at the basal zone of the leaves. The differentiation of vascular nodules always preceded bud formation. The difficulty of conifers to root and grow beyond plantlet stage is discussed. 相似文献
65.
Abstract Cyto-histological changes accompanying the formation of adventitious buds in excised cotyledons of Lactuca sativa were studied during the first 12 days after planting in vitro. Prospective proliferating cells can first be recognized, already on the first day after planting, by a marked increase in nuclear and nucleolar volumes, followed on the second day by a burst of cell divisions involving particularly mesophyll cells. Then lignified elements develop together with meristematic center, forming a callus-like tissue in the inner part of the cotyledons. At the third day of culture, the epidermal cells start to divide with a periclinal wall followed by an anticlinal division. In the following days of culture the epidermal cells, which divide mainly with periclinal walls, form layers of cells below the surface, gradually filling up the intercellular spaces. From the 8th day on, the buds protude above the surface and develops into shoots. These results are discussed in relation to DNA content of nuclei of Lactuca sativa cotyledons and to the time course of cell division and tracheary element formation. The very regular sequence of changes associated with the initiation and development of the bud makes the in vitro culture of Lactuca cotyledons an appropriate System for histochemical and biochemical studies. 相似文献
66.
Frederick C. Streich Jr. Virginia P. Ronchi J. Patrick Connick Arthur L. Haas 《The Journal of biological chemistry》2013,288(12):8209-8221
Ligation of polyubiquitin chains to proteins is a fundamental post-translational modification, often resulting in targeted degradation of conjugated proteins. Attachment of polyubiquitin chains requires the activities of an E1 activating enzyme, an E2 carrier protein, and an E3 ligase. The mechanism by which polyubiquitin chains are formed remains largely speculative, especially for RING-based ligases. The tripartite motif (TRIM) superfamily of ligases functions in many cellular processes including innate immunity, cellular localization, development and differentiation, signaling, and cancer progression. The present results show that TRIM ligases catalyze polyubiquitin chain formation in the absence of substrate, the rates of which can be used as a functional readout of enzyme function. Initial rate studies under biochemically defined conditions show that TRIM32 and TRIM25 are specific for the Ubc5 family of E2-conjugating proteins and, along with TRIM5α, exhibit cooperative kinetics with respect to Ubc5 concentration, with submicromolar [S]0.5 and Hill coefficients of 3–5, suggesting they possess multiple binding sites for their cognate E2-ubiquitin thioester. Mutation studies reveal a second, non-canonical binding site encompassing the C-terminal Ubc5α-helix. Polyubiquitin chain formation requires TRIM subunit oligomerization through the conserved coiled-coil domain, but can be partially replaced by fusing the catalytic domain to GST to promote dimerization. Other results suggest that TRIM32 assembles polyubiquitin chains as a Ubc5-linked thioester intermediate. These results represent the first detailed mechanistic study of TRIM ligase activity and provide a functional context for oligomerization observed in the superfamily. 相似文献
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Ronchi D Bordoni A Cosi A Rizzuti M Fassone E Di Fonzo A Servida M Sciacco M Collotta M Ronzoni M Lucchini V Mattioli M Moggio M Bresolin N Corti S Comi GP 《Biochemical and biophysical research communications》2011,(2):245-248
Leigh syndrome (LS) is an incurable, nearly always fatal, neurodegenerative, pediatric disorder that results from respiratory chain failure. The most common mitochondrial DNA (mtDNA) mutations that result in LS are m.8993T→C/G and m.9176T→C/G, which were previously found in several patients with early-onset Leigh syndrome. Here, we describe clinical and molecular features of a novel pedigree, where LS developed in two siblings. The proband was a young woman with an unusual adult-onset LS. She harbored a homoplasmic m.9176T→C mutation, based on analysis of a muscle biopsy. In contrast, the brother died at a young age. This novel case report and literature review highlights the variability of phenotypic expression of the m.9176T→C mutation. 相似文献
68.
Mitochondrial permeability transition is typically characterized by Ca2+ and oxidative stress-induced opening of a nonselective proteinaceous membrane pore sensitive to cyclosporin A, known as the
permeability transition pore (PTP). Data from our laboratory provide evidence that the PTP is formed when inner membrane proteins
aggregate as a result of disulfide cross-linking caused by thiol oxidation. Here we compared the redox properties between
PTP in intact mitochondria and mitoplasts. The rat liver mitoplasts retained less than 5% and 10% of the original outer membrane
markers monoamine oxidase and VDAC, respectively. Kidney mitoplasts also showed a partial depletion of hexokinase. In line
with the redox nature of the PTP, mitoplasts that were more susceptible to PTP opening than intact mitochondria showed higher
rates of H2O2 generation and decreased matrix NADPH-dependent antioxidant activity. Mitoplast PTP was also sensitive to the permeability
transition inducer tert-butyl hydroperoxide and to the inhibitors cyclosporin A, EGTA, ADP, dithiothreitol and catalase. Taken together, these data
indicate that, in mitoplasts, PTP exhibits redox regulatory characteristics similar to those described for intact mitochondria. 相似文献
69.
Daniel J. Edwards Frederick C. Streich Jr. Virginia P. Ronchi Dustin R. Todaro Arthur L. Haas 《The Journal of biological chemistry》2014,289(49):34114-34128
The human pathogen Shigella flexneri subverts host function and defenses by deploying a cohort of effector proteins via a type III secretion system. The IpaH family of 10 such effectors mimics ubiquitin ligases but bears no sequence or structural homology to their eukaryotic counterpoints. Using rates of 125I-polyubiquitin chain formation as a functional read out, IpaH9.8 displays V-type positive cooperativity with respect to varying concentrations of its Ubc5B∼125I-ubiquitin thioester co-substrate in the nanomolar range ([S]½ = 140 ± 32 nm; n = 1.8 ± 0.1) and cooperative substrate inhibition at micromolar concentrations ([S]½ = 740 ± 240 nm; n = 1.7 ± 0.2), requiring ordered binding to two functionally distinct sites per subunit. The isosteric substrate analog Ubc5BC85S-ubiquitin oxyester acts as a competitive inhibitor of wild-type Ubc5B∼125I-ubiquitin thioester (Ki = 117 ± 29 nm), whereas a Ubc5BC85A product analog shows noncompetitive inhibition (Ki = 2.2 ± 0.5 μm), consistent with the two-site model. Re-evaluation of a related IpaH3 crystal structure (PDB entry 3CVR) identifies a symmetric dimer consistent with the observed cooperativity. Genetic disruption of the predicted IpaH9.8 dimer interface reduces the solution molecular weight and significantly ablates the kcat but not [S]½ for polyubiquitin chain formation. Other studies demonstrate that cooperativity requires the N-terminal leucine-rich repeat-targeting domain and is transduced through Phe395. Additionally, these mechanistic features are conserved in a distantly related SspH2 Salmonella enterica ligase. Kinetic parallels between IpaH9.8 and the recently revised mechanism for E6AP/UBE3A (Ronchi, V. P., Klein, J. M., and Haas, A. L. (2013) E6AP/UBE3A ubiquitin ligase harbors two E2∼ubiquitin binding sites. J. Biol. Chem. 288, 10349–10360) suggest convergent evolution of the catalytic mechanisms for prokaryotic and eukaryotic ligases. 相似文献
70.
Anna Aspesi Elisa Pavesi Elisa Robotti Rossella Crescitelli Ilenia Boria Federica Avondo Hélène Moniz Lydie Da Costa Narla Mohandas Paola Roncaglia Ugo Ramenghi Antonella Ronchi Stefano Gustincich Simone Merlin Emilio Marengo Steven R. Ellis Antonia Follenzi Claudio Santoro Irma Dianzani 《Gene》2014
Defects in genes encoding ribosomal proteins cause Diamond Blackfan Anemia (DBA), a red cell aplasia often associated with physical abnormalities. Other bone marrow failure syndromes have been attributed to defects in ribosomal components but the link between erythropoiesis and the ribosome remains to be fully defined. Several lines of evidence suggest that defects in ribosome synthesis lead to “ribosomal stress” with p53 activation and either cell cycle arrest or induction of apoptosis. Pathways independent of p53 have also been proposed to play a role in DBA pathogenesis. 相似文献