Cavity spot of carrots: Interactions between the host and pathogen, related to the cell wall

This study investigates the structural aspects of cavity spot pathogenesis. Different Pythium spp. isolated from infected carrots, apples and melons were cultured on agar in Petri dishes and used for inoculation of uninfected carrots. Only slow-growing Pythium spp. (< 15 mm day^-1), such as P. violae and P. sulcatum caused cavity spot lesions. It is suggested that slow-growing species are able to penetrate, albeit slowly, into the plant tissue for 3 to 4 days before a hypersensitive reaction develops. Fast-growing species, however, did not cause lesions. Based on ultrastructural observations, we suggest that the following sequence of events occurs between the plant and the pathogen: The fungus infects the walls and grows for several days, during which time small amounts of wall-degrading enzymes are secreted. Phenylalanine ammonia lyase (PAL) activity and phenols increase linearly immediately upon inoculation. There was a lag phase of about 5 days before lignin began to increase linearly for about a month. Dissolution of wall components decreases the solute potential and water potential in the apoplast. Thus, water moves from the symplast into the apoplast, the turgor pressure gradually dissipates, and the cells shrink and eventually die.     

Water pathways in wheat leaves. III. The passage of the mestome sheath and the function of the suberised lamellae

The fluorochrome sulphorhodamine G, when present in the transpiration stream in wheat leaves, passes rapidly out of the veins and produces fluorescence in the mesophyll and epidermal cell walls. The path of movement of the dye out of the tracherary elements and across the mestome sheath to the parenchyma sheath cells was followed by rapid freezing, freeze-subsitution, dry embedding in resin, sectioning and epifluorescence microscopy. The sulphorhodamine solution was visible in tracheary elements, and, where it had passed out of the tracheary elements, strongly fluorescent in some of the cell walls. The patterns of wall fluorescence are used to chart the movements of water from the xylem through some of the radial walls of mestome sheath cells near the xylem to the free space of the mesophyll. The suberised lamellae of the mestome sheath cells must form an incomplete barrier near the xylem to permit passage of the dye. A hypothesis is formulated that the function of the suberised lamellae is to keep separate the oppositely directed fluxes of water and assimilates through the sheath. It is further proposed that the function of pits in living cells is a similar insulation of the symplastic traffic from the wayward waters of the apoplast.     

pH: Signal and Messenger in Plant Cells

Abstract: Since water spontaneously ionizes, protons cannot be removed from the medium: their free concentration in cells must be regulated through actively controlling H⁺‐related transport across membranes, by active and passive buffering, and by setting a certain pH within the metabolic network. Whereas these are the basic tools that provide effective H⁺ homeostasis, cellular compartmentation serves as an intermediate store into which protons can be shifted temporarily and from which protons can be regained when required. On the other hand, intracellular compartments can also serve as a final proton sink. pH regulation is not confined to intracellular spaces, but also comprises the apoplast. Whereas the pH of the cytosol is kept slightly alkaline at 7.2 to 7.5, with an average buffer capacity of 20 to 80 mM H⁺ per pH unit, the apoplastic pH may vary among tissues but is always acidic, with values between pH 5 and 6 and with a buffer capacity in the lower millimolar range per pH unit. pH can be a signal and/or a messenger, a distinction not always clearly made. Here, “signal” should be understood as information about an ongoing or preceding process, whereas “messenger” would be the carrying of certain information that will lead to a change of state. As such, pH would signal light intensity changes, drought, lack of oxygen and the presence of symbiotic partners or microbial attackers. On the other hand, pH would be a messenger in situations where pH changes are preconditions for certain processes, e.g., the gravity response or for activation of certain transporters in stomatal movements, and possibly for growth. The function of pH as a cellular messenger raises the question of whether pH should be understood as a “second messenger” in the way this is done for Ca²⁺. In an effort to give a comprehensive answer to this problem, the different roles of Ca²⁺ and H⁺ in cellular signalling are discussed and a number of Ca²⁺/pH interactions are presented.     

不同提取液提取水稻幼苗质外体蛋白效果的比较

提取植物组织质外体蛋白质的主要困难是提取效率低且易被细胞质蛋白污染。为解决上述问题,以12天93-11水稻幼苗为试验材料,使用3种含不同浓度钾和钙离子的缓冲液作为提取液进行提取效果比较。3种提取液的相同成分都是0.1mol/L Tris-HCl pH 7.6,1mmol/L PMSF,区别点在于:Buffer A含0.2mol/L KCl;Buffer B含0.2mol/L CaCl₂;Buffer C含0.1mol/L KCl和0.1mol/L CaCl₂。结果表明,Buffer A的蛋白产率达到了(0.49±0.07)mg/g FW(叶片)和(0.83±0.06)mg/g FW(根部),比Buffer B和Buffer C分别提高了122.7%和53.1%(叶片)以及102.4%和59.6% (根部)。六磷酸葡萄糖脱氢酶活性检测的结果表明在这些蛋白质提取物中细胞质蛋白的污染率很低,可以控制在1%以下。这些实验结果说明通过优化提取液,建立了有效提取水稻幼苗质外体蛋白质的方法,可应用于植物质外体蛋白质组学研究。     

Proteomics of ionically bound and soluble extracellular proteins in Medicago truncatula leaves

A large proportion of the apoplast proteome resides in the intercellular fluid (IF) or is ionically bound (IB) to the wall matrix. A combined analysis of IF and IB proteins of the Medicago truncatula leaf apoplast was performed. 2-DE analyses demonstrated the reproducible presence of 220 IF and 84 IB proteins in the apoplast. These two protein populations were largely distinct; 22 proteins could be spatially matched, but MALDI-TOF/TOF analyses suggested a considerably smaller number had common identities. MALDI-TOF/TOF characterisation identified 81 distinct proteins. Analyses of selected IF proteins (45) indicated 17 distinct proteins with mainly defence-related functions, whereas analyses of IB proteins (70) identified 63 distinct proteins of diverse natures, including proteins of non-canonical natures. The presence of non-canonical proteins in IB extracts is discussed in the light of evidence supporting a low level of contamination of purified walls from symplastic proteins. This work indicates that IB and IF proteins are functionally distinct fractions of the apoplast. The data obtained complements earlier studies of the Medicago proteome and therefore will be useful in future studies investigating the role of apoplastic proteins in plant processes.     

Changes in the lanthanum distribution following decapitation of the sporophytes of an aquatic fern,Marsilea drummondii A. Br.

Summary The active terminal bud and the quiescent lateral buds and corresponding nodes inserted at different levels on the main rhizome ofMarsilea drummondii were examined with the EM afterin vivo feeding with lanthanum nitrate. These tracer experiments demonstrate that all the buds are fed by their phloem cells. In the lateral bud axis the labelling of the sieve elements apoplast indicates that a solute transfer took place in the node between xylem and phloem via xylem transfer cells. La³⁺ deposits are completely absent from the apical dome of inhibited buds indicating that the walls of the quiescent meristematic cells are not permeated by the tracer. The removal of the terminal bud has two effects. It rapidly (in 2 hours) allows the lanthanum to penetrate the lateral bud tip walls at a stage when no fine structural changes are discernable and to bind to the outer surface of the plasmalemma as it does in the active terminal bud. This study including inhibited buds and buds released from apical dominance support the view that changes in the state of the cell surface (cell wall and plasma membrane) may be a prerequisite for the resumption growth activity.This study was supported in part by a grant from the Centre National de la Recherche Scientifique to L.Sossountzov (AI 031275).     

Plasmatubules in transfer cells of pea (Pisum sativum L.)

Plasmatubules are tubular evaginations of the plasmalemma. They have previously been found at sites where high solute flux between apoplast and symplast occurs for a short period and where wall proliferations of the transfer cell type have not been developed (Harris et al. 1982, Planta 156, 461–465). In this paper we describe the distribution of plasmatubules in transfer cells of the leaf minor veins of Pisum sativum L. Transfer cells are found in these veins associated both with phloem sieve elements and with xylem vessels. Plasmatubules were found, in both types of transfer cell and it is suggested that the specific distribution of the plasmatubules may reflect further membrane amplification within the transfer cell for uptake of solute from apoplast into symplast.     

Osmotic induction of fluid-phase endocytosis in onion epidermal cells

A transient plasmolysis/deplasmolysis (plasmolytic cycle) of onion epidermal cells has been shown to induce the formation of fluid-phase endocytic vesicles. Plasmolysis in the presence of the membrane-impermeant fluorescent probes Lucifer Yellow CH (LYCH) and Cascade Blue hydrazide resulted in the uptake of these probes by fluid-phase endocytosis. Following deplasmolysis, many of the dye-containing vesicles left their parietal positions within the cell and underwent vigorous streaming in the cytoplasm. Vesicles were observed to move within transvacuolar strands and their movements were recorded over several hours by video-microscopy. Within 2 h of deplasmolysis several of the larger endocytic vesicles had clustered around the nuclear membrane, apparently lodged in the narrow zone of cytoplams surrounding the nucleus. In further experiments LYCH was endocytically loaded into the cells during the first plasmolytic cycle and Cascade Blue subsequently loaded during a second plasmolytic cycle. This resulted in the introduction of two populations of endocytic vesicles into the cells, each containing a different probe. Both sets of vesicles underwent cytoplasmic streaming. The data are discussed in the light of previous observations of fluid-phase endocytosis in plant cells.     

Antioxidant defences of the apoplast

Summary The apoplast of barley and oat leaves contained superoxide dismutase (SOD), catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase activities. The activities of these enzymes in the apoplastic extracts were greatly modified 24 h after inoculation with the biotrophic fungal pathogenBlumeria graminis. The quantum efficiency of photosystem II, which is related to photosynthetic electron transport flux, was comparable in inoculated and healthy leaves during this period. Apoplastic soluble acid invertase activity was also modified in inoculated leaves. Inoculation-dependent increases in apoplastic SOD activity were observed in all lines. Major bands of SOD activity, observed in apoplastic protein extracts by activity staining of gels following isoelectric focusing, were similar to those observed in whole leaves but two additional minor bands were found in the apoplastic fraction. The apoplastic extracts contained substantial amounts of dehydroascorbate (DHA) but little or no glutathione (GSH). Biotic stress decreased apoplastic ascorbate and DHA but increased apoplastic GSH in resistant lines. The antioxidant cycle enzymes may function to remove apoplastic H₂O₂ with ascorbate and GSH derived from the cytoplasm. DHA and oxidized glutathione may be reduced in the apoplast or returned to the cytosol for rereduction.Abbreviations AA reduced ascorbate - APX ascorbate peroxidase - DHA dehydroascorbate (oxidised ascorbate) - DHAR dehydroascorbate reductase - G6PDH glucose-6-phosphate dehydrogenase - GSH reduced glutathione - GSSG glutathione disulphide - GR glutathione reductase - MDHA monodehydroascorbate - MDHAR monodehydroascorbate reductase - SOD superoxide dismutase