Biomedical subjects

Da-Peng Zhang

Publications and source records attributed to Da-Peng Zhang.

10 recordsLinked to original sources

PubMed2006

The Mg-chelatase H subunit is an abscisic acid receptor.

Abscisic acid (ABA) is a vital phytohormone that regulates mainly stomatal aperture and seed development, but ABA receptors involved in these processes have yet to be determined. We previously identified from broad bean an ABA-binding protein (ABAR) potentially involved in stomatal signalling, the gene for which encodes the H subunit of Mg-chelatase (CHLH), which is a key component in both chlorophyll biosynthesis and plastid-to-nucleus signalling. Here we show that Arabidopsis ABAR/CHLH specifically binds ABA, and mediates ABA signalling as a positive regulator in seed germination, post-germination growth and stomatal movement, showing that ABAR/CHLH is an ABA receptor. We show also that ABAR/CHLH is a ubiquitous protein expressed in both green and non-green tissues, indicating that it might be able to perceive the ABA signal at the whole-plant level.

Abscisic Acid↗

PubMed2006

A shift of Phloem unloading from symplasmic to apoplasmic pathway is involved in developmental onset of ripening in grape berry.

It remains unclear whether the phloem unloading pathway alters to adapt to developmental transition in fleshy fruits that accumulate high level of soluble sugars. Using a combination of electron microscopy, transport of the phloem-mobile symplasmic tracer carboxyfluorescein, movement of the companion cell-expressed and the green fluorescent protein-tagged viral movement protein, and assays of the sucrose cleavage enzymes, the pathway of phloem unloading was studied in the berries of a hybrid grape (Vitis vinifera x Vitis labrusca). Structural investigations showed that the sieve element-companion cell complex is apparently symplasmically connected through plasmodesmata with surrounding parenchyma cells throughout fruit development, though a small portion of plasmodesmata are apparently blocked in the ripening stage. Both carboxyfluorescein and the green fluorescent protein-tagged viral movement protein were released from the functional phloem strands during the early and middle stages of fruit development, whereas the two symplasmic tracers were confined to the phloem strands during the late stage. This reveals a shift of phloem unloading from symplasmic to apoplasmic pathway during fruit development. The turning point of the phloem unloading pathways was further shown to be at or just before onset of ripening, an important developmental checkpoint of grape berry. In addition, the levels of both the expression and activities of cell wall acid invertase increased around the onset of ripening and reached a high level in the late stage, providing further evidence for an operation of the apoplasmic unloading pathway after onset of ripening. These data demonstrate clearly the occurrence of an adaptive shift of phloem unloading pathway to developmental transition from growing phase to ripening in grape berry.

Carbohydrate Metabolism↗

PubMed2006

Abscisic acid stimulates a calcium-dependent protein kinase in grape berry.

It has been demonstrated that calcium plays a central role in mediating abscisic acid (ABA) signaling, but many of the Ca2+-binding sensory proteins as the components of the ABA-signaling pathway remain to be elucidated. Here we identified, characterized, and purified a 58-kD ABA-stimulated calcium-dependent protein kinase from the mesocarp of grape berries (Vitis vinifera x Vitis labrusca), designated ACPK1 (for ABA-stimulated calcium-dependent protein kinase1). ABA stimulates ACPK1 in a dose-dependent manner, and the ACPK1 expression and enzyme activities alter accordantly with the endogenous ABA concentrations during fruit development. The ABA-induced ACPK1 stimulation appears to be transient with a rapid effect in 15 min but also with a slow and steady state of induction after 60 min. ABA acts on ACPK1 indirectly and dependently on in vivo state of the tissues. Two inactive ABA isomers, (-)-2-cis, 4-trans-ABA and 2-trans, 4-trans-(+/-)-ABA, are ineffective for inducing ACPK1 stimulation, revealing that the ABA-induced effect is stereo specific to physiological active (+)-2-cis, 4-trans-ABA. The other phytohormones such as auxin indoleacetic acid, gibberellic acid, synthetic cytokinin N-benzyl-6-aminopurine, and brassinolide are also ineffective in this ACPK1 stimulation. Based on sequencing of the two-dimensional electrophoresis-purified ACPK1, we cloned the ACPK1 gene. The ACPK1 is expressed specifically in grape berry covering a fleshy portion and seeds, and in a developmental stage-dependent manner. We further showed that ACPK1 is localized in both plasma membranes and chloroplasts/plastids and positively regulates plasma membrane H+-ATPase in vitro, suggesting that ACPK1 may be involved in the ABA-signaling pathway.

Abscisic Acid↗

PubMed2004

Evidence for apoplasmic phloem unloading in developing apple fruit.

The phloem unloading pathway remains unclear in fleshy fruits accumulating a high level of soluble sugars. A structural investigation in apple fruit (Malus domestica Borkh. cv Golden Delicious) showed that the sieve element-companion cell complex of the sepal bundles feeding the fruit flesh is symplasmically isolated over fruit development. 14C-autoradiography indicated that the phloem of the sepal bundles was functional for unloading. Confocal laser scanning microscopy imaging of carboxyfluorescein unloading showed that the dye remained confined to the phloem strands of the sepal bundles from the basal to the apical region of the fruit. A 52-kD putative monosaccharide transporter was immunolocalized predominantly in the plasma membrane of both the sieve elements and parenchyma cells and its amount increased during fruit development. A 90-kD plasma membrane H(+)-ATPase was also localized in the plasma membrane of the sieve element-companion cell complex. Studies of [14C]sorbitol unloading suggested that an energy-driven monosaccharide transporter may be functional in phloem unloading. These data provide clear evidence for an apoplasmic phloem unloading pathway in apple fruit and give information on the structural and molecular features involved in this process.

Biological Transport↗

PubMed2004

Membrane-associated protein kinase activities in the developing mesocarp of grape berry.

Fruit development is a process involving various signals and gene expression. Protein phosphorylation catalyzed by protein kinases is known to play a key role in eukaryotic cell signalling and so may be involved in the regulation of fruit development. Using the method of exogenous substrate phosphorylation, we characterised the calcium-dependent and calmodulin-independent protein kinase (CDPK) activity and the myelin basic protein (MBP)-phosphoralating activity that could be due to a mitogen-activated protein kinase (MAPK)-like activity in the developing mesocarp of grape berry. The CDPK activity was shown to be predominantly localised in the plasma membrane, while the MAPK-like activity was predominantly associated with endomembranes. The assays of bivalent cation requirement showed that Mn2+ could to a certain extent replace Mg2+ in the incubation system for the protein kinase activities. Both CDPK and MAPK-like activities were resistant to heat treatment. The activities of the two enzymes were fruit developmental stage-specific with the highest activities of both enzymes in the lag growth phase before the ripening stage, suggesting strongly the important roles of the detected CDPK and MAPK-like activities in the fruit development.

Calcium↗

PubMed2004

Abscisic acid is involved in the water stress-induced betaine accumulation in pear leaves.

ABA exogenously applied to the leaves of the whole plants of pear (Pyrus bretschneideri Redh. cv. Suly grafted on Pyrus betulaefolia Rehd.) significantly increased the betaine concentrations in the leaves when the plants were well watered. The plants subjected to 'drought plus ABA' treatment had significantly higher betaine concentrations in their leaves than those given drought treatment alone. The 'drought plus ABA' treatment increased the amount of betaine aldehyde dehydrogenase (BADH, EC 1.2.1.8) and its activity in the leaves more than did the drought treatment alone. The experiments with detached leaves showed that ABA treatment significantly increased the concentration of betaine, activity of BADH and apparent amount of BADH in non-dehydrated leaves, and enhanced the accumulation of betaine, activity of BADH and apparent amount of BADH in dehydrated leaves. These effects of ABA were both time- and dose-dependent. Two ABA isomers, (-)-cis, trans-ABA and 2-trans, 4-trans-ABA, had no effect on the betaine accumulation in the leaves, showing that the ABA-induced effects are specific. These data demonstrate that ABA is involved in the drought-induced betaine accumulation in the pear leaves.

Abscisic Acid↗

PubMed2004

Phloem unloading in developing walnut fruit is symplasmic in the seed pericarp and apoplasmic in the fleshy pericarp.

The sieve element-companion cell (SE-CC) complex of the sepal bundles feeding the fleshy pericarp of developing walnut (Juglans regia L.) fruit is structurally symplasmically isolated, but the SE-CC complex of the minor ventral carpellary bundles located in the seed pericarp and feeding the seed is structurally symplasmically connected to its adjacent parenchyma cells. 14C-autoradiography indicated that the phloem of both the sepal and carpellary bundles was functional for unloading. Confocal laser scanning microscopy imaging of carboxyfluorescein unloading showed that the dye is confined to the phloem strands of the sepal bundles in the fleshy pericarp, but released from the phloem strands of the minor ventral carpellary bundles into the surrounding parenchyma cells in the seed pericarp. A 60-kDa acid invertase was immunolocalized to the cell wall of SE-CC complex and parenchyma cells in both the fleshy and seed pericarp. These data provide clear evidence for an apoplasmic phloem unloading pathway in the fleshy pericarp and a predominant symplasmic phloem unloading pathway parallel with a possible apoplasmic path as suggested by the presence of the extracellular invertase in the seed pericarp. A model of complex phloem unloading pathways in developing walnut fruit has been proposed.

Carbon Radioisotopes↗

PubMed2003

Membrane-associated protein kinase activities in developing apple fruit.

Fruit development is a process involving various signals and gene expression. Protein phosphorylation catalysed by protein kinases is known to play a key role in eukaryotic cell signalling and so may be involved in the regulation of fruit development. Using the method of exogenous substrate phosphorylation, the activity of calcium-dependent and calmodulin-independent protein kinase (CDPK) that was stimulated by phosphatidylserine, and the myelin basic protein (MBP)-phosphorylating activity that could be due to a calcium-independent mitogen-activated protein kinase-like (MAPK-like) activity in the developing apple fruits were identified. The CDPK activity was shown to be predominantly localized in the plasma membrane, whereas in the presence of phosphatidylserine, the high activity of CDPK was detected in both plasma membrane and endomembranes. The MAPK-like activity was predominantly associated with endomembranes. The assays of bivalent cation requirement showed that Mn2+ could replace Mg2+ in the incubation system for the protein kinase activities and stimulate CDPK activity more than Mg2+. Heat treatment abolished CDPK but stimulated MAPK-like activity. The activities of the phosphatidylserine-stimulated CDPK and of the MAPK-like were fruit developmental stage-specific with higher activities of both enzymes in the early and middle developmental stages in comparison with the late developmental stage. These data suggest that the detected protein kinases may play an important role in the fruit development.

Journal Article↗

PubMed2002

Purification and identification of a 42-kilodalton abscisic acid-specific-binding protein from epidermis of broad bean leaves.

Purification of abscisic acid (ABA)-binding proteins is considered to constitute a major step toward isolating ABA receptors. We report here that an ABA-binding protein was for the first time, to our knowledge, purified from the epidermis of broad bean (Vicia faba) leaves via affinity chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, isoelectric focusing electrophoresis, and isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis two-dimensional electrophoresis of the purified ABA-binding protein all identified a single protein band with a molecular mass of 42 kD and an isoelectric point 4.86. The Scatchard plot for the purified protein showed a linear function with a maximum binding activity of 0.87 mol mol(-1) protein and an equilibrium dissociation constant of 21 nM, indicating that the purified protein may be a monomeric one, possessing one binding site. The ABA-binding protein was enriched more than 300-fold with a yield of 14%. (-)ABA and trans-ABA were substantially incapable of displacing (3)H-(+/-)ABA bound to the ABA-binding protein, and (+/-)ABA was less effective than (+)ABA in the competition. These findings allow establishment of the stereospecificity of the 42-kD protein and suggest its ABA receptor nature. Pretreatment of the guard cell protoplasts of broad bean leaves with the monoclonal antibody raised against the 42-kD protein significantly decreased the ABA specific-induced phospholipase D activity in a dose-dependent manner. This physiological significance provides more clear evidence for the potential ABA-receptor nature of the 42-kD protein.

Abscisic Acid↗

PubMed2002

[Study on optimization of expression, purification, properties and biological function of recombinant human sBLyS].

The prokaryotic expression plasmid pET-30a(+)/sBLyS was constructed and transformed into E. coli BL21 (lambda DE3). The recombinant protein was found to be highly expressed by the plight of soluble part and inclusion body. For the sake of enhancing the proportion of the soluble part, inducement at 16 degrees C for 12 h was ascertained. The expressing product was then purified by Ni2+ affinity chromatography gel. PI of the recombinant human sBLyS(rhsBLyS) is about 7.1-7.3 and it assembles into a homotrimer. The effect of rhsBLyS on B lymphocytes by MTT method told us the B lymphocytes' proliferating capacity dose depended on concentration and also stimulating time of the rhsBLys. With rhsBLyS(2 micrograms/mL) stimulating 3 days, B lymphocytes can proliferate the most.

B-Cell Activating Factor↗