Aggregation-related association of lipid with the cytoskeleton of rabbit and human platelets prelabeled with [3H]palmitic acid. Similar effects of adenosine diphosphate- and thrombin-induced aggregation.

Livné, Avinoam; Packham, MA; Guccione, MA; Mustard, J. F.

doi:10.1172/jci113320

Cited by 15 publications

(5 citation statements)

References 37 publications

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“…Extensive crosslinking of the cytoskeleton and membrane skeleton mdy occur, leading to pelleting of part of the membrane skeleton with the cytoskeletal pellet. For instance, the appearance of lipids in the cytoskeleton of thrombin-activated platelets has been re-ported [35], and in the present study the second phase of myosin incorporation in the cytoskeleton coincided with its virtual disappearance from the membrane skeleton. Membrane skeleton and cytoskeleton may therefore have different meanings in non-activated versus activated cells.…”

Section: Discussionsupporting

confidence: 74%

Vinculin is a permanent component of the membrane skeleton and is incorporated into the (re)organising cytoskeleton upon platelet activation

Asijee

Sturk

Bruin

et al. 1990

European Journal of Biochemistry

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Vinculin, a 130-kDa protein discovered in chicken gizzard smooth-muscle cells and subsequently also described in platelets, is believed to be involved in membrane-cytoskeleton interactions. In this study we investigated vinculin distribution in human blood platelets. Two skeletal fractions and a remaining cytosolic fraction were prepared with a recently described Triton X-100 lysis buffer causing minimal post-lysis breakdown by proteolysis. The presence of vinculin was demonstrated in the membrane skeleton and cytosol of resting and thrombin-activated human platelets. Upon thrombin stimulation vinculin also appeared in the cytoskeleton. This cytoskeletal incorporation was completed during the early stages of platelet aggregation and secretion, when the uptake of myosin, actin-binding protein and talin was still not maximal. We conclude therefore, that vinculin may play an important role in the structural (re)organisation of the human platelet cytoskeleton upon platelet activation.Vinculin is a 130-kDa protein which was first discovered in chicken gizzard smooth-muscle cells [ 11, and subsequently also in other cells [2 -51, including bovine [6] and human blood platelets 17 -101. There are two main lines of evidence which suggest that vinculin is a member of the protein complex that links actin filaments to the plasma membrane, and is thus involved in cytoskeleton-membrane attachment [I -41. First, in several cell types the protein was found to be located at specialized sites where microfilament bundles terminate close to the plasma membrane: the dense plaques of smooth-muscle cells [3, We recently demonstrated the incorporation of vinculin into the cytoskeleton of bovine blood platelets upon thrombin stimulation [15]. However, there were two main reasons for investigating this incorporation further. First, the incorporation was much slower than the platelet aggregation reaction, which would indicate a role for vinculin solely in the later stages of platelet activation. Second, platelets contain not only a cytoskeleton but also a membrane skeleton, i.e. a peripheral layer of actin filaments, composed of short filaments crosslinked by actin-binding protein and linked to the plasma membrane, primarily to the glycoprotein Ib/IX complex [16]. This layer lies just below the lipid bilayer and appears to follow the contours of the plasma membrane. It remains at the periphery

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Section: Discussionsupporting

confidence: 74%

Vinculin is a permanent component of the membrane skeleton and is incorporated into the (re)organising cytoskeleton upon platelet activation

Asijee

Sturk

Bruin

et al. 1990

European Journal of Biochemistry

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“…Results are the means + SEM of three to five independent experiments. of PtdInsP2-containing, Triton X-100-resistant membrane fragments in the cytoskeleton of large platelet aggregates (Zucker and Masiello, 1983;Livne et al, 1988) we measured the total cytoskeletal lipid (measured as lipid phosphate) under different conditions. Although the amount of phospholipid associated with the cytoskeleton increases -10-fold in platelet aggregates compared with unstimulated cells, it does so with an entirely different (faster) time course than that of the increase in cytoskeletal PtdInsP2 levels (not shown).…”

Section: Resultsmentioning

confidence: 99%

Aggregation-dependent, integrin-mediated increases in cytoskeletally associated PtdInsP2 (4,5) levels in human platelets are controlled by translocation of PtdIns 4-P 5-kinase C to the cytoskeleton.

1996

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Thrombin‐stimulated aggregation of human platelets promotes an increase in the phosphatidylinositol 4‐phosphate (PtdIns 4‐P) 5‐kinase (PIPkin) activity in the cytoskeleton. This phenomenon is associated with translocation of PIPkin isoform C to the cytoskeleton and with an increase in the amount of phosphatidylinositol bisphosphate (PtdInsP2) bound to the cytoskeletal pellet. All three of these effects are prevented if the platelets are not stirred or if RGD‐containing peptides are present, demonstrating that they require integrin activation. All three are also abolished by pretreatment with okadaic acid, which also prevents the aggregation‐dependent translocation of pp60(c‐src) to the cytoskeleton. The results point to the existence of a cytoskeletally associated PtdInsP2 pool under the control of integrin‐mediated signals that act via PIPkin C and suggest that a common, okadaic acid‐sensitive mechanism may underlie the aggregation‐dependent translocation of certain signalling molecules to the platelet cytoskeleton.

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“…Electron microscopic studies indicate that portions of the platelet membrane are not lysed when Triton X-100 is added to strongly aggregated platelets [6,27]. These unlysed fragments are probably responsible for the presence of some of the '*'I-labeled membrane glycoproteins [6] and [3H]palmitate-labeled phospholipids [27] that are found in the Triton-insoluble residue of aggregated platelets several minutes after aggregation is initiated. The presence of such membrane fragments may account for the increase in the amount of protein and radiolabeled glycoproteins that we noted between 3 and 6 min after ADP-induced aggregation.…”

mentioning

confidence: 99%

“…The presence of such membrane fragments may account for the increase in the amount of protein and radiolabeled glycoproteins that we noted between 3 and 6 min after ADP-induced aggregation. Others have shown that when Triton was added when aggregation was only 10-20% complete, the [3H]phospholipid that became Triton-insoluble was probably not present in unlysed membrane fragments, since it was released if actin was depolymerized [27]. However, under those conditions, GP IIIa was not incorporated into the cytoskeletal core [27].…”

mentioning

confidence: 99%

“…Others have shown that when Triton was added when aggregation was only 10-20% complete, the [3H]phospholipid that became Triton-insoluble was probably not present in unlysed membrane fragments, since it was released if actin was depolymerized [27]. However, under those conditions, GP IIIa was not incorporated into the cytoskeletal core [27]. The study of Wheeler et al [7] probably provides the best evidence in the literature that membrane glycoproteins can be specifically incorporated into the cytoskeletal core and that GP IIb/IIIa were the only labeled proteins incorporated when Triton was added as soon as aggregation reached its peak.…”

mentioning

confidence: 99%

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Effect of the thiol group inhibitor monobromobimane and other inhibitors on the composition of the platelet cytoskeletal core and its association with glycoprotein IIIa

Puszkin

Mauss

Milot

et al. 1989

J of Cellular Biochemistry

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SDS-polyacrylamide gel electrophoresis was used to study the effects of the thiol inhibitor monobromobimane (MB), EDTA, and prostaglandin E1 (PGE1) on the formation and composition of the platelet cytoskeletal core (Triton-insoluble residue) and its association with glycoprotein (GP) IIIa. Stimulation or aggregation of platelets in response to ADP or thrombin increased the amount of Triton-insoluble myosin. Aggregation resulted in incorporation of [125I]GP IIIa and a new band at about 210 kDa into the cytoskeletal core. EDTA and PGE1 caused little disaggregation of platelets that were aggregated in PRP with ADP and that had secreted the contents of their granules. In contrast to EDTA, PGE1 decreased the amount of Triton-insoluble residue and its association with GP IIIa. MB added after ADP-induced aggregation caused an increase in the amount of cytoskeletal core despite marked disaggregation and a substantial decrease in core-associated GP IIIa. With aspirin-treated platelets that had not secreted, EDTA, PGE1, and MB all caused disaggregation and loss of cytoskeletal GP IIIa. MB diminished, but did not reverse, thrombin-induced aggregation of washed platelets and arrested GP IIIa incorporation into the cytoskeletal core. Concanavalin A (Con A) cross-links glycoproteins on a single platelet and induces incorporation of GP IIIa into the Triton-insoluble residue in the absence of platelet aggregation. This induction was not inhibited by MB, although this reagent, as well as aspirin, inhibited Con A-induced secretion. Since GP IIIa incorporation caused by ADP-induced aggregation differs from that caused by Con A in its susceptibility to MB, it seems unlikely that thiol groups are directly involved in the association of GP IIIa with the cytoskeletal core.

show abstract

Aggregation-related association of lipid with the cytoskeleton of rabbit and human platelets prelabeled with [3H]palmitic acid. Similar effects of adenosine diphosphate- and thrombin-induced aggregation.

Cited by 15 publications

References 37 publications

Vinculin is a permanent component of the membrane skeleton and is incorporated into the (re)organising cytoskeleton upon platelet activation

Vinculin is a permanent component of the membrane skeleton and is incorporated into the (re)organising cytoskeleton upon platelet activation

Aggregation-dependent, integrin-mediated increases in cytoskeletally associated PtdInsP2 (4,5) levels in human platelets are controlled by translocation of PtdIns 4-P 5-kinase C to the cytoskeleton.

Effect of the thiol group inhibitor monobromobimane and other inhibitors on the composition of the platelet cytoskeletal core and its association with glycoprotein IIIa

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