Inhibitory action of polyamines on protein kinase C association to membranes

Moruzzi, Maria Stella; Barbiroli, Bruno; Monti, Maria Giuseppina; Tadolini, Bruna; Hakim, Gabriele; Mezzetti, Gabriele

doi:10.1042/bj2470175

Cited by 49 publications

(25 citation statements)

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“…Furthermore, Ca2+ appears to be more specific than Mg2+. A similar competition between polyamines and calcium for binding to PM was earlier observed by Moruzzi et al (1987) in human erythrocyte membrane vesicles, where it was proposed to represent a mechanism to modulate protein kinase C activation. On the other hand, the inhibition of spermidine binding by calcium may also reflect an activation of proteases.…”

Section: Dlscusslonsupporting

confidence: 71%

Polyamine Binding to Plasma Membrane Vesicles Isolated from Zucchini Hypocotyls

Tassoni¹,

Antognoni²,

Bagni³

1996

Plant Physiol.

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Aliphatic polyamines are ubiquitous polycationic substances in prokaryotic and eukaryotic cells that influence a variety of cellular processes, although their mechanism of action remains to be clarified. In plants they are implicated in the regulation of developmental processes such as embryogenesis, senescence, and flowering.Because of their protonated amino and imino groups at physiological pH values, polyamines bind to anionic cell constituents such as nucleic acids, phospholipids, negatively charged protein residues of membranes, and pectic substances of cell walls. In vivo and in vitro interactions with DNA, tRNA, and rRNA are well known (Bagni et al., 1982;Igarashi et al., 1982;Feuerstein and Marton, 1989). Evidence suggests that their role is to stabilize and affect the conformation of nucleic acids and their activities, resulting in improved translation and protein synthesis (Cocucci and Bagni, 1968;Quigley et al., 1978).The interaction between polyamines and membranes is suggested to be an intermediate in many important cellular events such as membrane fusion (Schuber et al., 1983) transmission of receptor-mediated signals (Koenig et al., 1983), and regulation of prostaglandin synthesis (Igarashi et al., 1981). Nevertheless, information on interactions between polyamines and the PM is very scarce, although their ability to stabilize biological membranes has been demonstrated (Schuber, 1989). Studies in vitro have shown that polyamines can bind to artificial membranes such as liposomes and that the binding strength of the complexes depends on the type and density of acidic phospholipids present in the vesicles (Tadolini et al., 1986).In animals, specific polyamine-binding proteins have been detected in severa1 tissues and body fluids. A spermine-binding protein was found, for example, in the cytosol of duodenal mucosa of newborn chick (Mezzetti et al., 1981). This interaction can be regulated by the hormonal form of vitamin D, (Mezzetti et al., 1983).Because systematic studies have yet to be undertaken on this topic in plants, in the present work we have attempted to define the characteristics of spermidine binding to purified PM isolated from etiolated hypocotyls of zucchini (Cucurbita pepo L.).The experiments reported here represent a preliminary step toward the identification of putative polyaminebinding sites and/or carriers at the plasmalemma leve1 in plant cells. MATERIALS A N D METHODS Plant MaterialSeeds of zucchini (Cucurbita pepo L. hybrid Storr's Green, Asgrow Co., Lodi, Italy) were planted in moist vermiculite and grown at 22 ? 1°C in total darkness for 7 d. Etiolated hypocotyls (6-12 cm in length) were cut off and stored at -20°C until use. Preparation and Characterization of Membrane VesiclesA11 procedures were carried out on ice under room light.The plant material (60-80 g of hypocotyls) was homogenized in a mortar using 2 volumes of buffer A containing 50 mM Tris-HC1, pH 7.5, 2.0 mM EDTA, 250 mM SUC, and 10 mM 2-mercaptoethanol. The homogenate was filtered through four layers of cheeseclo...

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Section: Dlscusslonsupporting

confidence: 71%

Polyamine Binding to Plasma Membrane Vesicles Isolated from Zucchini Hypocotyls

Tassoni¹,

Antognoni²,

Bagni³

1996

Plant Physiol.

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“…Phorbol esters, which activate protein kinase C, inhibit the L-type channel current in this preparation (Linden & Routtenberg, 1989). Putrescine is known to inhibit protein kinase C but does not affect the cyclic AMP-dependent protein kinase (Qi, Schatzman, Mazzei, Turner, Raynor, Liao & Kuo, 1983;Thams, Capito & Hedeskov, 1986;Moruzzi, Barbiroli, Monti, Tadolini, Hakim & Mezzetti, 1987). It was therefore hypothesized that the effect of putrescine might be mediated via protein kinase C.…”

Section: Discussionmentioning

confidence: 99%

The effect of polyamines on voltage‐activated calcium channels in mouse neuroblastoma cells.

Herman

Reuveny

Narahashi

1993

The Journal of Physiology

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SUMMARY1. Putrescine has been implicated in modulating cytoplasmic calcium concentration and is correlated with selective neuronal vulnerability in cerebral ischaemia. In order to determine whether putrescine modulates voltage-activated calcium channels, whole-cell and single channel patch clamp experiments were performed with NIE-115 mouse neuroblastoma cells.2. L-type calcium channel currents showed a 34 + 21 % increase (n = 6 cells) during external application of 1 mm putrescine. There was no change in the kinetics of the current and no shift in the current-voltage relationship along the voltage axis.3. T-type calcium channel currents were not affected by 1 mM putrescine. 4. The effect of putrescine on single L-type calcium channels was studied using the cell-attached configuration of the patch clamp technique. Putrescine (5 mM) applied to the bathing solution, but not present in the pipette, caused an increase in open time of the single channel current without changing the conductance of the channel. In 345 depolarizing steps compiled from three cells, the number of channel openings longer than 3 ms increased from six to seventy-six, and the number of channel openings longer than 9 ms increased from zero to twenty-seven. This single channel study supports the hypothesis that putrescine acts on the L-type channel from the inside of the cell.5. External application of 1 mm spermine and 1 mm spermidine had no effect on T-and L-type calcium channels. Thus, the effect of putrescine is probably not mediated by the higher polyamines.6. In order to test whether the effect of putrescine is mediated by a second messenger, specific protein kinase C and cyclic AMP-dependent protein kinase inhibitors, staurosporine and KT5720, respectively, were applied prior to putrescine. When cells were preconditioned with 200 nm staurosporine, the increase of the Ltype calcium current by 1 mm putrescine was inhibited. By contrast, 200 nM KT5720 did not inhibit the putrescine effect. Therefore, the increase of L-type channel currents by putrescine may be mediated by protein kinase C but not the cyclic AMPdependent protein kinase. MS 1220 M. D. HERMAN, E. REUVENY AND T. NARAHASHI 7. The putrescine-induced enhancement of the L-type calcium channel activity may play an important role in calcium-induced neurotoxicity.

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“…This was borne out in model systems. Moruzzi et al (1987), using protein kinase C purified from rat brain and inside-out human erythrocyte membrane vesicles, were able to show that micromolar concentrations of spermine (spermidine was much less potent) greatly interfered with the formation of the active membraneassociated enzyme complex. That this might not be the only explanation for the inhibitory action of polyamines on protein kinase C activity was verified by the same group, which showed that spermine at millimolar concentrations interacted also with the catalytic domain of the enzyme and strongly inhibits its phosphorylation activity (Mezzetti et al, 1988).…”

Section: Polyamines and Transportmentioning

confidence: 99%