Expression of scinderin, an actin filament‐severing protein, in different tissues

Tchakarov, L.; Vitale, María Leiza; Jeyapragasan, M.; Castillo, A. Rodríguez Del; Trifaró, J. M.

doi:10.1016/0014-5793(90)81010-l

Cited by 52 publications

(33 citation statements)

References 16 publications

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“…These results do not rule out the possibility that other actin-binding proteins might have stronger affinities for actin. However, the present results as well as our earlier published experiments (Rodrfguez Del Castillo et al, 1990) demonstrate that both gelsolin and scinderin are equally retained by actin affinity columns.…”

Section: Ca 2+ and Ph Dependent Interactions Of Scinderin With Phosphsupporting

confidence: 82%

“…Control experiments were carried out with (a) FITC-IgG alone and (b) scinderin antiserum previously adsorbed with scinderin. Scinderin antibodies were raised in rabbits against the purified bovine protein (Rodrfguez Del Castillo et al, 1990), scinderin antiserum 6 has been previously characterized (Tchakarov et ai., 1990;Vitaie et al, 1991).…”

Section: Immunofluorescence Microscopy Studiesmentioning

confidence: 99%

“…Homogenates were centrifuged at 100,000 g for 1 h, sediments were discarded and supernatants (cytosolic protein fraction) were used. Scinderin was purified from bovine adrenal medullae as previously described (Rodrfguez Del Castillo et al, 1990). Cytosolic protein fractions and scinderin preparations were dialyzed overnight at 4~ against buffer B or C. Phosphatidylinositol 4,5-biphosphate (sodium salt, PIP2) was dissolved in buffer B or C at a concentration of I mg/ml.…”

Section: Binding Studiesmentioning

confidence: 99%

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Ca2+ and pH determine the interaction of chromaffin cell scinderin with phosphatidylserine and phosphatidylinositol 4,5,-biphosphate and its cellular distribution during nicotinic-receptor stimulation and protein kinase C activation.

Castillo

Vitale

Trifaró

1992

The Journal of Cell Biology

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Abstract. Nicotinic stimulation and high K § -ization of chromaffin cells cause disassembly of cortical filamentous actin networks and redistribution of scinderin, a Ca2+-dependent actin filament-severing protein. These events which are Ca2+-dependent precede exocytosis. Activation of scinderin by Ca ~ § may cause disassembly of actin filaments leaving cortical areas of low cytoplasmic viscosity which are the sites of exocytosis (Vitale, M. L., A. Rodriguez Del Castillo, L. Tchakarov, and J.-M. Trifar6. 1991. J. Cell. Biol. 113:1057-1067. It has been suggested that protein kinase C (PKC) regulates secretion. Therefore, the possibility that PKC activation might modulate scinderin redistribution was investigated. Here we report that PMA, a PKC activator, caused scinderin redistribution, although with a slower onset than that induced by nicotine. PMA effects were independent of either extra or intracellular Ca 2 § as indicated by measurements of Ca 2 § transients, and they were likely to be mediated through direct activation of PKC because inhibitors of the enzyme completely blocked the response to PMA. Scinderin was not phosphorylated by the kinase and further experiments using the Na+/H § antiport inhibitors and intracellular pH determinations, demonstrated that PKC-mediated scinderin redistribution was a consequence of an increase in intracellular pH. Moreover, it was shown that scinderin binds to phosphatidylserine and phosphatidylinositol 4,5-biphosphate liposomes in a Ca2 § manner, an effect which was modulated by the pH. The results suggest that under resting conditions, cortical scinderin is bound to plasma membrane phospholipids. The results also show that during nicotinic receptor stimulation both a rise in intracellular Ca 2 § and pH are observed. The rise in intracellular pH might be the result of the translocation and activation of PKC produced by Ca ~ § entry. This also would explain why scinderin redistribution induced by nicotine is partially (26-40%) inhibited by inhibitors of either PKC or the Na+/H § antiport. In view of these findings, a model which can explain how scinderin redistribution and activity may be regulated by pH and Ca 2 § in resting and stimulated conditions is proposed.

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Section: Ca 2+ and Ph Dependent Interactions Of Scinderin With Phosphsupporting

confidence: 82%

Section: Immunofluorescence Microscopy Studiesmentioning

confidence: 99%

Section: Binding Studiesmentioning

confidence: 99%

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Ca2+ and pH determine the interaction of chromaffin cell scinderin with phosphatidylserine and phosphatidylinositol 4,5,-biphosphate and its cellular distribution during nicotinic-receptor stimulation and protein kinase C activation.

Castillo

Vitale

Trifaró

1992

The Journal of Cell Biology

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“…Previously, bovine adseverin had only been purified from adrenal and exocytotic cells (31). It was also immunologically detected in pituitary, brain, and kidney tissues after partial enrichment (25,28,37), again pointing at the low abundance of the protein. It is important to mention here that mouse adseverin is strongly upregulated in mast cells, which contain many granules and are known to be exocytotically active.…”

Section: Discussionmentioning

confidence: 98%

Murine Adseverin (D5), a Novel Member of the Gelsolin Family, and Murine Adseverin Are Induced by Interleukin-9 in T-Helper Lymphocytes

Robbens

Louahed

Pestel

et al. 1998

Molecular and Cellular Biology

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We identified a number of upregulated genes by differential screening of interleukin-9-stimulated T-helper lymphocytes. Interestingly, two of these messengers encode proteins that are similar to proteins of the gelsolin family. The first displays a typical structure of six homologous domains and shows a high level of identity (90%) with bovine adseverin (or scinderin) and may therefore be considered the murine adseverin homolog. The second encodes a protein with only five segments. Sequence comparison shows that most of the fifth segment and a short amino-terminal part of the sixth segment (amino acids 528 to 628 of adseverin) are missing, and thus, this form may represent an alternatively spliced product derived from the same gene. The corresponding protein is called mouse adseverin (D5). We expressed both proteins in Escherichia coli and show that mouse adseverin displays the typical characteristics of all members of the gelsolin family with respect to actin binding (capping, severing, and nucleation) and its regulation by Ca 2؉ . In contrast, mouse adseverin (D5) fails to nucleate actin polymerization, although like mouse adseverin and gelsolin, it severs and caps actin filaments in a Ca 2؉ -dependent manner. Adseverin is present in all of the tissues and most of the cell lines tested, although at low concentrations. Mouse adseverin (D5) was found only in blood cells and in cell lines derived from T-helper lymphocytes and mast cells, where it is weakly expressed. In a gel filtration experiment, we demonstrated that mouse adseverin forms a 1:2 complex with G actin which is stable only in the presence of Ca 2؉ , while no stable complex was observed for mouse adseverin (D5

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“…To study the distribution of scinderin and gelsolin in bovine adrenal chromaflin cells in culture, it was important to demonstrate the specificity of the polyclonal antisera to be used. Previous work from our laboratory demonstrated that scinderin and gelsolin antisera are specific for these proteins and that do not cross-react with any other chromaffin cell cytosolic protein (Bader et al, 1986;Rodriguez Del Castillo et al, 1990;Tchakarov et al, 1990). Cross-reactivity of both antisera with chromaffin cell cytoskeletal proteins was also investigated.…”

Section: R E S U L T S (A) Specificity Of Scinderin and Gelsolin Antimentioning

confidence: 95%

Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.

Vitale

Castillo

Tchakarov

et al. 1991

The Journal of Cell Biology

202

191

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Abstract.Immunofluorescence and cytochemical studies have demonstrated that filamentous actin is mainly localized in the cortical surface of the chromaffin cell. It has been suggested that these actin filament networks act as a barrier to the secretory granules, impeding their contact with the plasma membrane. Stimulation of chromaffin cells produces a disassembly of actin filament networks, implying the removal of the barrier. The presence of gelsolin and scinderin, two Ca2+-dependent actin filament severing proteins, in the cortical surface of the chromatfin cells, suggests the possibility that cell stimulation brings about activation of one or more actin filament severing proteins with the consequent disruption of actin networks. Therefore, biochemical studies and fluorescence microscopy experiments with scinderin and gelsolin antibodies and rhodamine-phalloidin, a probe for filamentous actin, were performed in cultured chromaffin cells to study the distribution of scinderin, gelsolin, and filamentous actin during cell stimulation and to correlate the possible changes with catecholamine secretion. Here we report that during nicotinic stimulation or K+-evoked depolarization, subcortical scinderin but not gelsolin is redistributed and that this redistribution precedes catecholamine secretion. The rearrangement of scinderin in patches is mediated by nicotinic receptors. Cell stimulation produces similar patterns of distribution of scinderin and filamentous actin. However, after the removal of the stimulus, the recovery of scinderin cortical pattern of distribution is faster than F-actin reassembly, suggesting that scinderin is bound in the cortical region of the cell to a component other than F-actin. We also demonstrate that peripheral actin filament disassembly and subplasmalemmal scinderin redistribution are calcium-dependent events. Moreover, experiments with an antibody against dopamine-~-hydroxylase suggest that exocytosis sites are preferentially localized to areas of F-actin disassembly.

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Expression of scinderin, an actin filament‐severing protein, in different tissues

Cited by 52 publications

References 16 publications

Ca2+ and pH determine the interaction of chromaffin cell scinderin with phosphatidylserine and phosphatidylinositol 4,5,-biphosphate and its cellular distribution during nicotinic-receptor stimulation and protein kinase C activation.

Ca2+ and pH determine the interaction of chromaffin cell scinderin with phosphatidylserine and phosphatidylinositol 4,5,-biphosphate and its cellular distribution during nicotinic-receptor stimulation and protein kinase C activation.

Murine Adseverin (D5), a Novel Member of the Gelsolin Family, and Murine Adseverin Are Induced by Interleukin-9 in T-Helper Lymphocytes

Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.

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