Interaction of ruthenium red with Ca2+-binding proteins

Aphids, which constitute one of the most important groups of agricultural pests, ingest nutrients from sieve tubes, the photoassimilate transport conduits in plants. Aphids are able to successfully puncture sieve tubes with their piercing mouthparts (stylets) and ingest phloem sap without eliciting the sieve tubes' normal occlusion response to injury. Occlusion mechanisms are calciumtriggered and may be prevented by chemical constituents in aphid saliva injected into sieve tubes before and during feeding. We recorded aphid feeding behavior with the electrical penetration graph (EPG) technique and then experimentally induced sieve tube plugging. Initiation of sieve tube occlusion caused a change in aphid behavior from phloem sap ingestion to secretion of watery saliva. Direct proof of ''unplugging'' properties of aphid saliva was provided by the effect of aphid saliva on forisomes. Forisomes are proteinaceous inclusions in sieve tubes of legumes that show calcium-regulated changes in conformation between a contracted state (below calcium threshold) that does not occlude the sieve tubes and a dispersed state (above calcium threshold) that occludes the sieve tubes. We demonstrated in vitro that aphid saliva induces dispersed forisomes to revert back to the nonplugging contracted state. Labeling Western-blotted saliva proteins with 45 Ca 2؉ or ruthenium red inferred the presence of calcium-binding domains. These results demonstrate that aphid saliva has the ability to prevent sieve tube plugging by molecular interactions between salivary proteins and calcium. This provides aphids with access to a continuous flow of phloem sap and is a critical adaptation instrumental in the evolutionary success of aphids.calcium-binding ͉ plant-aphid interaction ͉ plugging ͉ saliva proteins ͉ sieve element

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“…3b). The same proteins were stained on a nitrocellulose membrane by using ruthenium red (34) (Fig. 3c).…”

Section: Resultsmentioning

confidence: 99%

Molecular sabotage of plant defense by aphid saliva

Will

Tjallingii

Thönnessen

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

455

413

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“…RuR is known to interact with several Ca 2 þ -binding proteins [13][14][15][16] and to inhibit Ca 2 þ transport in mitochondria, 17 VDAC channel activity and PTP opening. 7,10,12,17 Purified recombinant native and E72Q-mVDAC1 were reconstituted into a planar lipid bilayer (PLB) as a single or multichannel and the currents produced in response to voltages stepped from a holding potential of 0 to À20 mV were recorded ( Figure 3).…”

Section: Rur Induces Channel Closure Of Native But Not E72q-mutated Mmentioning

confidence: 99%

“…7,10,12 Ruthenium red (RuR), a hexavalent polysaccharide stain, is known to interact specifically with Ca 2 þ -binding proteins. 7,10,[13][14][15][16] RuR has been shown to inhibit several mechanisms involved in intracellular Ca 2 þ regulation, including the mitochondrial uniporter and the 'rapid mode' system, 17 sarcoplasmic reticulum Ca 2 þ -ATPase activity, 13 Ca 2 þ release and ryanodine binding by the Ca 2 þ release channel/ryanodine receptor, 14 the binding of Ca 2 þ to calsequestrin 15 and to calmodulin, 16 as well as to prevent brain tubulin polymerization. 18 Recently, we have shown that RuR and the trivalent cation Ru360, a derivative of RuR, 19 can induce VDAC channel closure in a time-dependent manner, stabilizing the channel in a completely closed state.…”

Section: Introductionmentioning

confidence: 99%

The voltage-dependent anion channel-1 modulates apoptotic cell death

et al. 2005

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The role of the voltage-dependent anion channel (VDAC) in cell death was investigated using the expression of native and mutated murine VDAC1 in U-937 cells and VDAC inhibitors. Glutamate 72 in VDAC1, shown previously to bind dicyclohexylcarbodiimide (DCCD), which inhibits hexokinase isoform I (HK-I) binding to mitochondria, was mutated to glutamine. Binding of HK-I to mitochondria expressing E72Q-mVDAC1, as compared to native VDAC1, was decreased by B70% and rendered insensitive to DCCD. HK-I and ruthenium red (RuR) reduced the VDAC1 conductance but not that of E72Q-mVDAC1. Overexpression of native or E72Q-mVDAC1 in U-937 cells induced apoptotic cell death (80%). RuR or overexpression of HK-I prevented this apoptosis in cells expressing native but not E72Q-mVDAC1. Thus, a single amino-acid mutation in VDAC prevented HK-I-or RuR-mediated protection against apoptosis, suggesting the direct VDAC regulation of the mitochondria-mediated apoptotic pathway and that the protective effects of RuR and HK-I rely on their binding to VDAC.

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“…In tobacco seedlings, ruthenium red prevents any increase in cytosolic [Ca"] othenvise caused by wind-induced motion (Knight et al, 1992). Additionally, in animal cells ruthenium red competitively binds to Ca2+-binding proteins (Charuk et al, 1990), including CM, with inhibition of CaCMdependent enzyme activity (Sasaki et al, 1992). In roots of maize seedlings and in maize suspension-cultured cells, ruthenium red blocked the increase in cytosolic [Ca2+] that would have occurred when cells were made anoxic (Subbaiah et al, 1994a, 199413).…”

Section: Cellulase Activity and Cell Deathmentioning

confidence: 99%

Transduction of an Ethylene Signal Is Required for Cell Death and Lysis in the Root Cortex of Maize during Aerenchyma Formation Induced by Hypoxia

1996

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Ethylene has been implicated in signaling cell death in the lysigenous formation of gas spaces (aerenchyma) in the cortex of adventitious roots of maize (Zea mays) subjected to hypoxia. Various antagonists that are known to modify particular steps in signal transduction in other plant systems were applied at low concentrations to normoxic and hypoxic roots of maize, and the effect on cell death (aerenchyma formation) and the increase in cellulase activity that precedes the appearance of cell degeneration were measured. 60th cellulase activity and cell death were inhibited in hypoxic roots in the presence of antagonists of inositol phospholipids, CaZ+-calmodulin, and protein kinases. By contrast, there was a parallel promotion of cellulase activity and cell death in hypoxic and normoxic roots by contact with reagents that activate G-proteins, increase cytosolic CaZ+, or inhibit protein phosphatases. Most of these reagents had no effect on ethylene biosynthesis and did not arrest root extension. These results indicate that the transduction of an ethylene signal leading to an increase in intracellular Ca2+ is necessary for cell death and the resulting aerenchyma development in roots of maize subjected to hypoxia.

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Interaction of ruthenium red with Ca2+-binding proteins

Cited by 132 publications

References 58 publications

Molecular sabotage of plant defense by aphid saliva

Molecular sabotage of plant defense by aphid saliva

The voltage-dependent anion channel-1 modulates apoptotic cell death

Transduction of an Ethylene Signal Is Required for Cell Death and Lysis in the Root Cortex of Maize during Aerenchyma Formation Induced by Hypoxia

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