Plasma membrane phosphorylation by endogenous phosphate donors in human blood platelets Selectivity of the action of dibutyryl cyclic AMP

Apitz‐Castro, Rafael; Ramirez, E.; Maingón, R.; Murciano, Alicia de; Ribbi, Alida

doi:10.1016/0005-2736(76)90312-6

Cited by 20 publications

(9 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The existence of a phosphoglycoprotein with an apparent molecular weight greater than 400,000 dalton (HMW-GP) in human platelets plasma membranes has been clearly demonstrated in our laboratory (5,6), and a similar component has also been described by Nurden et al (7) and McMaster et al (8). The later authors showed that it can be digested by bovine thrombin, although the conditions used were much more drastic than required for platelets to undergo thrombin-induced aggregation.…”

supporting

confidence: 69%

Inhibition of the Platelet Reaction by a High Molecular Weight Phosphoglycoprotein Isolated from Human Platelet Plasma Membranes

et al. 1981

View full text Add to dashboard Cite

SummaryThe effect of a phospho-glycoprotein (HMW-GP), obtained from human platelet plasma membranes, on the aggregation and secretion of human platelets was studied. Incubation of PRP with 4 to 16 μg/ml of HMW-GP results in inhibition of ADP-, Epinephrine-, Collagen-, and Thrombin-induced platelet aggregation. The effect is mainly reflected on the secondary wave of aggregation. The inhibitory effect is partially overcome by higher concentration of the inducers, however, even under these conditions, a clear tendency towards disaggregation is observed. 5HT release (Col-induced) is strongly decreased from 50% to 4.5. The inhibitory effect on Thrombin-induced aggregation is markedly dependent on external calcium, being maximal at 5 mM calcium. The HMW-GP does not bind ADP or Thrombin. Membrane conformation is markedly affected, as evidenced by the effect of HMW-GP on the iodination of surface polypeptides of intact platelets. It is suggested that interaction of HMW-GP with the platelet membrane blocks the signal(s) transmission that links stimulus to activation. The inhibition observed might just represent an experimental amplification of the endogenous modulatory function that has been proposed for this high molecular weight phosphoglycoprotein.

show abstract

supporting

confidence: 69%

Inhibition of the Platelet Reaction by a High Molecular Weight Phosphoglycoprotein Isolated from Human Platelet Plasma Membranes

et al. 1981

View full text Add to dashboard Cite

show abstract

“…The increase of cAMP levels in platelets produced the phosphorylation of several proteins with molecular masses from 20 to 250 kDa (13)(14)(15)(16)(17)(18)(19)(20). Two of those proteins have estimated molecular masses of 22 and 24 kDa, and they are rapidly phosphorylated by cAMP-dependent protein kinase (15,16).…”

Section: -106mentioning

confidence: 99%

A ras-related protein is phosphorylated and translocated by agonists that increase cAMP levels in human platelets.

Lapetina

Lacal

Reep

et al. 1989

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

132

View full text Add to dashboard Cite

The antigenicity of platelet proteins was assayed against various monoclonal antibodies (mAbs) that recognize specific epitopes of the ras-encoded p21 protein. mAb M90, which detects the region of p21 protein within amino acids 107-130 and inhibits its GTP-binding activity, strongly reacted with a 22-kDa protein present in the particulate fraction of human platelets. Other mAbs against ras-encoded proteins, including Y13-259, which efficiently detects ras proteins from a variety of organisms, did not recognize the platelet 22-kDa protein. Transfer ofthe platelet 22-kDa protein to nitrocellulose paper showed that the protein binds [a-32P]GTP. Moreover, preincubation of the transferred protein with mAb M90 drastically reduced its GTP-binding activity. Treatment of platelets with ioprost, a prostacyclin analog, caused (i) a time-dependent increase of a 24-kDa protein that is recognized by mAb M90 in particulate and cytosolic fractions and (ii) the gradual decrease of the 22-kDa protein from the particulate fraction. When platelets were labeled with 32P and then treated with iloprost, the 24-kDa protein was found to be phosphorylated. The 32P-labeled 24-kDa protein was specifically immunoprecipitated by mAb M90. These results suggest that appearance of the 24-kDa protein results from phosphorylation of the 22-kDa protein, which shifts its mobility to a higher molecular mass area.In the past two years platelets have been shown to possess distinct GTP-binding proteins with molecular masses between 21 and 31 kDa (1-3). These low molecular weight guanine nucleotide-binding regulatory (G) proteins can be electrophoretically transferred from SDS/polyacrylamide gels to nitrocellulose blots, where they are detected by the binding of [a-32P]GTP (2, 3). One of them, a 21-kDa protein, was recently isolated from platelet membranes; however, this protein is not recognized with monoclonal antibody (mAb) Y13-259 (4), which recognizes all known ras-encoded p21 proteins (1). Also, this antibody did not cross-react with the other low molecular mass G proteins present in platelets (3). The possible physiological significance of these proteins has not yet been elucidated; they could be correlated with regulation of phospholipase C (2) and, recently, we reported that a platelet ras-related protein is phosphorylated by a cAMP-dependent protein kinase (5).Lacal and Aaronson (6) have developed a series of mAbs that recognize epitopes located at different regions in the Harvey (Ha) ras-encoded p21 protein. We have screened platelet proteins against those mAbs and found that a 22-kDa platelet protein is recognized by two of those mAbs. These mAbs, designated M3 and M90, recognize a region of ras p21 protein containing residues 116-119, involved in the interaction with the guanine base (7). By contrast the 22-kDa protein was not recognized by the ras-specific mAb Y13-259. These results suggest that the 22-kDa platelet protein is a member ofthe ras family. Finally, the 22-kDa protein is a substrate for cAMP-dependent protein kinase. Ph...

show abstract

“…The supernatant plasma was carefully collected in a transfer bag (Fenwall TA-3M) without anticoagulant. In all cases the residual erythrocytes were removed from the platelet-rich plasma by repeating the centrifugation at 700g for 55s as previously described (Apitz-Castro et al, 1976). Plateletpoor plasma was obtained by centrifugation of platelet-rich plasma at 3000g for 10min, followed by 30min centrifugation of the supernatant plasma at 30000g.…”

Section: Methodsmentioning

confidence: 99%

“…A Gilford 240 spectrophotometer (Gilford Instruments Laboratories, Oberlin, OH, U.S.A.), whichwas modified to include a magnetic stirrer under the cell compartment, was used to measure the extent of platelet aggregation as described previously (Apitz-Castro et al, 1976). In experiments designed to study the effect of colchicine on the degree of platelet aggregation, a saturation curve for ADP was always established beforehand, in order to select the concentration of ADP giving the maximum and halfmaximum effects.…”

Section: Platelet Aggregationmentioning

confidence: 99%

“…Phosphorylation of tubulin-like material from intact platelets suspended in plasma Platelet-rich plasma (40ml) was incubated for 1 h with 150pCi Of [32P]P,/ml in 0.15 pH7.3,at 37°C, as described previously by Apitz-Castro et al (1976). For the experiments designed to study the effect of colchicine on the phosphorylation of tubulin, platelet-rich plasma was divided into two equal samples (after addition of the [32p]p;), and colchicine was added to one of them at a final concentration of 2mM.…”

Section: Platelet Aggregationmentioning

confidence: 99%

See 1 more Smart Citation

The effect of colchicine on human blood platelets under conditions of short-term incubation

Ribbi-Jaffe¹,

Apitz‐Castro²

1979

Biochemical Journal

View full text Add to dashboard Cite

The effects of colchicine on ADP-induced aggregation and on the phosphorylation of tubulin-like protein from human blood platelets were studied. Colchicine at 2mM concentration completely inhibits ADP-induced aggregation after 8min incubation. Under the same inhibitory conditions, phosphorylation of tubulin-like materials in intact platelets was also impaired whereas the endogenous kinase activity of tubulin, isolated through polymerization--depolymerization cycles, was not affected. It was also shown that, under conditions of maximal inhibition of both aggregation and tubulin phosphorylation, colchicine does not penetrate into the cells. The results obtained suggest that the effect of colchicine on platelet aggregation might be mainly, although not exclusively, due to a non-specific effect of the alkaloid on the plasma membrane, rather than to a direct action of the drug on the microtubular protein subunits.

show abstract

Plasma membrane phosphorylation by endogenous phosphate donors in human blood platelets Selectivity of the action of dibutyryl cyclic AMP

Cited by 20 publications

References 28 publications

Inhibition of the Platelet Reaction by a High Molecular Weight Phosphoglycoprotein Isolated from Human Platelet Plasma Membranes

Inhibition of the Platelet Reaction by a High Molecular Weight Phosphoglycoprotein Isolated from Human Platelet Plasma Membranes

A ras-related protein is phosphorylated and translocated by agonists that increase cAMP levels in human platelets.

The effect of colchicine on human blood platelets under conditions of short-term incubation

Contact Info

Product

Resources

About