Accumulation of potentiometric and other dyes in haustoria of Erysiphe graminis in living host cell

Bushnell, W. R.; Mendgen, Kurt; Liu, Z.

doi:10.1016/0885-5765(87)90068-3

Cited by 14 publications

(6 citation statements)

References 26 publications

(27 reference statements)

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“…6A). This resembled transient peaks obtained with haustorial mitochondria (4) and probably represented a period in which dye was released from mitochondria but before the dye diffused out of the cell.…”

supporting

confidence: 68%

Pontentiometric Cyanine Dyes Are Sensitive Probes for Mitochondria in Intact Plant Cells

Liu¹,

Bushnell²,

Brambl³

1987

Plant Physiol.

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Selected fluorescent dyes were tested for uptake by mitochrondria in intact cells of barley, maize, and onion. The cationic cyanine dye 3,3'-diheptyloxacarbocyanine iodide [DiOC7(3)j accumulated in mitochondria within 15 to 30 minutes without appreciable staining of other protoplasmic constituents. The number, shape, and movement of the fluorescent mitochondria could be seen readily, and the fluorescence intensity of the mitochondria could be monitored with a microscope photometer. Fluorescence was eliminated in 1 to 5 minutes by the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) indicating that maintenance of dye concentration was dependent on the inside-negative transmembrane potential maintained by functional mitochondria. Fluorescence of prestained mitochondria was enhc within 5 to 10 minutes after addition of 0.1 minlimolar kinetin to cells. The fluorescence in kinetintreated cells was dissipated by CCCP. These results suggest that kinetin interacted with respiratory processes resulting in higher potential across the mitochondrial membrane.Fluorescent probes have been applied as optical indicators of membrane potential differences in several types of cells, isolated organelles, and lipid vesicles (2,6,29,30). The technique relies on potential-dependent partitioning of charged lipophillic dye molecules across the membrane. Changes in membrane potential result in changes in intensity of dye fluorescence, termed 'redistribution signals' (6).Potentiometric dyes have been used successfully to measure changes in membrane potential of mitochondria within or isolated from cells of yeasts (15,24), several kinds of animal cells (3,7,8,13,14,16,25), and recently, in cells or protoplasts of higher plants (18,26 (30) and co-workers for use as potentiometric probes. Through the use ofrespiratory inhibitors and ionophores and ofothers means, the fluorescence intensity of these dyes has been shown to be related to mitochondrial membrane potential (2, 6, 8, 13.).We have recently found that certain cyanine dyes gave fluorescence to mitochondria of haustoria within living barley epidermal cells infected with Erysiphe graminis (4). The mitochondrial fluorescence was rapidly dissipated by the protonophores CCCP2 or DNP indicating that maintenance of dye concentration within the mitochondria was dependent on membrane potential. Several of the cyanines tested were observed also to stain mitochondria of host epidermal cells, which prompted us to screen various cyanines and other dyes used elsewhere as mitochondrial probes for ability to stain higher plant mitochondria. Our objectives were to learn which dyes were most effective with cells of higher plants, to learn if fluorescence of stained plant mitochondria could be monitored photometrically (as had been possible for haustorial mitochondria), and to learn if fluorescence was related to mitochondrial membrane potential. We also measured the effect of kinetin on mitochondrial fluorescence. Our interest in kinetin grew out of the fact that kinetin had interf...

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supporting

confidence: 68%

Pontentiometric Cyanine Dyes Are Sensitive Probes for Mitochondria in Intact Plant Cells

Liu¹,

Bushnell²,

Brambl³

1987

Plant Physiol.

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“…hordei as reported earlier (Bushnell et al 1987). There was only a small increase in fluorescence of the host cytoplasm including mitochondria (Fig.…”

Section: Resultssupporting

confidence: 84%

“…This procedure was performed with the whole mount positioned under the microscope objective lens. Compared to our earlier measurements (Bushnell et al 1987), the method was improved to perform an exchange of the fluids within 30 s. For measurements of fluorescence intensity, a microscope photometer (MPV 2; Leitz, Wetzlar, FRG) equipped with an incident-illumination system (Ploemopak with a 100-W, DC, highpressure mercury lamp) and a filter block I2 (BP 45~490, RKP 510, LP 515) was used with a Leitz fluorescence objective 40/0.75. The measuring diaphragm had an opening of 9" 3 gm and was focused exactly over the center of the haustorial body with the mitochondria.…”

Section: Methodsmentioning

confidence: 49%

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The activity of powdery-mildew haustoria after feeding the host cells with different sugars, as measured with a potentiometric cyanine dye

Mendgen

Nass

1988

Planta

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Abstract. The biotrophic parasite Erysiphe graminisf. sp. hordei produces haustoria within the cells of its host Hordeum vulgare. To determine the physiological activity of these haustoria, the electric potential across the membranes in the mitochondria of the haustorium was studied. The membrane potential was estimated with the fluorescent potentiometric cyanine dye 3,3'-dibutyloxacarbocyanine iodide. The addition of depolarizing agents (carbonylcyanide m-chlorophenylhydrazone, 2,4-dinitrophenol or KCN) to infected cells resulted in an increase of fluorescence after the addition of low concentrations or a decrease of fluorescence after the addition of higher concentrations. When the infected host cell was fed with increasing concentrations of D-glucose (25, 50, 75 raM), corresponding decreases of fluorescence were measured immediately in the mitochondria of the fungal haustoria. Sucrose induced a similar reduction of fluorescence about 20 min later. D-Galactose and D-fructose induced a somewhat smaller reduction of fluorescence, L-glucose and D-glucitol had no effect. The results indicate that haustoria take up glucose from the host cells immediately. Sucrose, D-galactose and D-fructose seem to require time to be metabolized before their products reach the fungal haustorium or mitochondria.

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“…hordei), we have evidence now that glucose is the sugar preferably transported from host to parasite. To show this, the membrane potential of haustorial mitochondria was studied with a potentiometric cyanine dye (BUSHNELL et al 1987) which exhibits fluorescence in a polarized membrane. We assumed that the sugar which reaches the haustorium would first increase respiration there.…”

Section: Time (Minutes)mentioning

confidence: 99%

The Differentiation of Infection Structures as a Result of Recognition Events between some Biotrophic Parasites and their Hosts

Mendgen

Schneider

Sterk

et al. 1988

Journal of Phytopathology

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MOSl uredospores of rUSl fungi develop infeclion SlrUClUres in a lypical pallern so lhal lhey can infecl lhe hosl plam. The funclion of lhese infeclion SlrUClures is divided intO lhe following three phases:I. In lhe recognilion phase, lhe germ lube recognizes lhe cUlicle and lhe stOma. This process may occur independemly from lhe hOSl plant since copies of lhe cUlicle induce similar reaclions of lhe fungus. During fungal growlh on lhe epidermis, unspecific Slress responses of lhe planl are lriggered. . 2. In lhe signal phase, lhe fungal substOmalal vesicle and infeclion hypha(e) 'contacl lhe hOSl cells wilhin lhe leaf parenchyma. A signal from lhe hosl induces funher developmem of lhe fungus. HaustOrium mOlher cell differentialion is effecled and hauslorium formalion is inilialed. Al lhe same lime, lhe fungus suppresses lhe synlhesis of slfe;s melaboliles by lhe planl. 3. In lhe parasilic phase, lhe fungus penelrales lhe hOSl cell and complex imeraclions belween hOSl and parasile begin. A highly specialized interface around lhe haustOrium develops presumably in order to allow a more efficient nUlriem lransfer from hOSl IQ parasile. Eventual defence reaclions of lhe plant, generally on lhe race-cuilivar level, fail IQ be evoked or are suppressed in compalible combinalions.

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Accumulation of potentiometric and other dyes in haustoria of Erysiphe graminis in living host cell

Cited by 14 publications

References 26 publications

Pontentiometric Cyanine Dyes Are Sensitive Probes for Mitochondria in Intact Plant Cells

Pontentiometric Cyanine Dyes Are Sensitive Probes for Mitochondria in Intact Plant Cells

The activity of powdery-mildew haustoria after feeding the host cells with different sugars, as measured with a potentiometric cyanine dye

The Differentiation of Infection Structures as a Result of Recognition Events between some Biotrophic Parasites and their Hosts

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