Ca2+ concentration during binding determines the manner in which annexin V binds to membranes

Trotter, Patrick J.; Orchard, Margaret A.; Walker, John H.

doi:10.1042/bj3080591

Cited by 63 publications

(48 citation statements)

References 57 publications

(55 reference statements)

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“…Such proteins have already been proposed to explain the interaction of annexin with cell membranes at low Ca# + concentrations [29] and might provide an explanation for the ability of annexin V to inhibit cPLA # activity in permeabilized HL-60 cells [30].…”

Section: Discussionmentioning

confidence: 97%

Inhibition of human cytosolic phospholipase A2 by human annexin V

Buckland

Wilton

1998

Biochemical Journal

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The ability of annexins, particularly annexin 1 (lipocortin 1), to inhibit phospholipase A2 (PLA2) is well known and a substrate depletion mechanism is now widely accepted as the explanation for most inhibitory studies. However, there are only a very limited number of reported studies involving annexins and the high-molecular-mass cytosolic PLA2 (cPLA2). In this study we have examined the effect of human recombinant annexin V, a potentially abundant cytosolic protein, on the ability of human recombinant cPLA2 to hydrolyse a variety of phospholipid substrates. The results show clearly that, under the conditions of our study, annexin V can inhibit cPLA2 activity by a mechanism of substrate depletion and that this inhibition is dependent on the nature of the phospholipids and the concentration of Ca2+ ions in the assay. The hydrolysis of 1-stearoyl 2-arachidonyl phosphatidylcholine by cPLA2 was not significantly affected by annexin V over a range of Ca2+ concentrations (1 μM-2.5 mM), a result that presumably reflects the zwitterionic nature of the phospholipid and the known inability of annexins to bind to such interfaces. In contrast, the hydrolysis of dioleoyl phosphatidylglycerol, which is an effective anionic phospholipid substrate for this enzyme, and more significantly that of 1-stearoyl 2-arachidonyl phosphatidic acid, were readily inhibited by annexin V, although these effects were Ca2+-dependent. The Ca2+ concentrations required for inhibition in the assay system in vitro are greater than those associated with Ca2+-stimulated events within the cell, suggesting that a role for annexin V in regulating cPLA2 activity might not involve a substrate depletion mechanism in vivo unless factors in addition to Ca2+ and phospholipids contribute to the binding of annexin V to cell membranes.

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Section: Discussionmentioning

confidence: 97%

Inhibition of human cytosolic phospholipase A2 by human annexin V

Buckland

Wilton

1998

Biochemical Journal

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“…At a concentration that allows cPLA 2 membrane translocation (0.8 M) (35), a recent study on platelets has demonstrated that annexin V is firmly bound to membranes (51). A major part of membrane-bound annexin V (85%) is resistant to EGTA extraction that could be due to a still unidentified membrane protein.…”

Section: Discussionmentioning

confidence: 99%

“…The inhibitory activity of annexin V was suppressed by heating the protein at 100°C for 5 min, confirming that this activity depends on the integrity of the structure. ] i concentration in the micromolar range can physiologically occur in subcellular compartment after cell activation (50) and has been reported to be required for annexin V binding to artificial phospholipid vesicles (51).…”

Section: Release Of [ 3 H]arachidonic Acid From Slo-permeabilized Hl-mentioning

confidence: 99%

Inhibition of Cytosolic Phospholipase A2 by Annexin V in Differentiated Permeabilized HL-60 Cells

Mira

Dubois

Oudinet

et al. 1997

Journal of Biological Chemistry

107

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Annexin V belongs to a family of proteins that interact with phospholipids in a Ca 2؉ -dependent manner. This protein has been demonstrated to have anti-phospholipase A 2 activity. However, this effect has never yet been reported with the 85-kDa cytosolic PLA 2 (cPLA 2 ). We studied, in a model of differentiated and streptolysin O-permeabilized HL-60 cells, the effect of annexin V on cPLA 2 activity after stimulation by calcium, GTP␥S (guanosine 5-O-(3-thiotriphosphate)), formyl-Met-LeuPhe, or phorbol 12-myristate 13-acetate. Both recombinant and human placental purified annexin V inhibit cPLA 2 activity whatever the stimulus used. The decrease of arachidonic acid release is of 40 and 50%, respectively, at [Ca 2؉ ] of 3 and 10 M. The mechanism of inhibition was also analyzed. cPLA 2 requires calcium and protein kinase C (PKC) or mitogen-activated protein kinase phosphorylation for its activation. As annexin V was shown to be an endogenous inhibitor of PKC, PKC-stimulated cPLA 2 activity was analyzed. Using GF109203x, a specific PKC inhibitor, we demonstrated that this pathway is of minor importance in our model. cPLA 2 inhibition by annexin V is not linked to PKC inhibition. To test the hypothesis of phospholipid depletion, mutants of annexin V were constructed using mutagenesis directed to Ca 2؉ site. We demonstrate that the Ca 2؉ site located in domain I is necessary for the inhibitory effect of annexin V on cPLA 2 activity. The site in domain IV is also involved but with less efficiency. In contrast, mutations in site II and III do not modify this effect. Moreover, annexin V mutated on all sites does not inhibit cPLA 2 . Thus, we propose a predominant role of module (I/IV) in the biological action of annexin V, which, in physiological conditions, may control cPLA 2 activity by depletion of the phospholipid substrate.

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“…Because it was known that annexins could be partially extracted by EGTAcontaining solution (Trotter et al, 1995), we tested whether the presence or absence of Ca 2ϩ during isolation of CPP might modify the yield of annexins (Fig. 2).…”

Section: Quantification Of Annexins II V and Vi Proteins In Human Mmentioning

confidence: 99%

Expression and Localization of the Annexins II, V, and VI in Myocardium from Patients with End-Stage Heart Failure

Benevolensky

Belikova

Mohammadzadeh

et al. 2000

Laboratory Investigation

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SUMMARY:Annexins II, V, and VI belong to a family of Ca 2ϩ -dependent phospholipid-binding proteins that have been involved mainly in signal transduction, differentiation, membrane trafficking events, or binding to the extracellular matrix, or that might be effective as Ca 2ϩ -channels. They are abundant in the mammalian myocardium and might play a role in ventricular remodeling and altered calcium handling during heart failure. To test this hypothesis, we compared the expression and distribution of these annexins in nonfailing (n ϭ 9) and failing human hearts with idiopathic dilated cardiomyopathy (n ϭ 11). Northern blot and slot blot analysis were used to determine the annexin mRNA levels and Western blots were used to quantify the amounts of annexin proteins. Distribution of annexins was studied by immunohistofluorescence labeling and compared with that of a sarcolemmal marker (Na ϩ /K ϩ -ATPase) and of a myofibrillar protein (␣-actinin). We showed that nonfailing hearts contained a higher amount of annexin VI than of annexin V or II (13.5 Ϯ 1.8, 3.7 Ϯ 0.2, and 2.5 Ϯ 0.5 g/mg protein, respectively). In failing hearts, there was a parallel increase in both mRNA and protein levels of annexin II (146% and 132%, p Ͻ 0.05, respectively) and annexin V (152%, p Ͻ 0.01, 147%, p Ͻ 0.005, respectively); the protein level of annexin VI was also increased (117%, p Ͻ 0.05), whereas the increase of its mRNA level was statistically insignificant. We observed a predominant localization of annexin II in interstitium, and of annexins V and VI in cardiomyocytes at the level of the sarcolemma, T-tubules, and intercalated disks in nonfailing hearts, whereas in failing hearts enlarged interstitium contained all three annexins. Furthermore, annexin V staining at the level of cardiomyocytes almost disappeared. In conclusion, we showed that heart failure is accompanied by marked overexpression of annexins II and V, as well as translocation of annexin V from cardiomyocytes to interstitial tissue. The data suggest that annexins may contribute to ventricular remodeling and annexin V to impaired Ca 2ϩ handling in failing heart. (Lab Invest 2000, 80:123-133).

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Ca2+ concentration during binding determines the manner in which annexin V binds to membranes

Cited by 63 publications

References 57 publications

Inhibition of human cytosolic phospholipase A2 by human annexin V

Inhibition of human cytosolic phospholipase A2 by human annexin V

Inhibition of Cytosolic Phospholipase A2 by Annexin V in Differentiated Permeabilized HL-60 Cells

Expression and Localization of the Annexins II, V, and VI in Myocardium from Patients with End-Stage Heart Failure

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