SIGNAL AMPLIFICATION IN HL-60 GRANULOCYTES - EVIDENCE THAT THE CHEMOTACTIC PEPTIDE RECEPTOR CATALYTICALLY ACTIVATES GUANINE-NUCLEOTIDE-BINDING REGULATORY PROTEINS IN NATIVE PLASMA-MEMBRANES

被引:94
作者
GIERSCHIK, P
MOGHTADER, R
STRAUB, C
DIETERICH, K
JAKOBS, KH
机构
[1] Pharmakologisches Institut, Universität Heidelberg
来源
EUROPEAN JOURNAL OF BIOCHEMISTRY | 1991年 / 197卷 / 03期
关键词
D O I
10.1111/j.1432-1033.1991.tb15964.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Receptors for the chemotactic peptide fMet-Leu-Phe (fMet, N-formylmethionine) are present in membranes of myeloid differentiated human leukemia (HL-60) cells and stimulate phospholipase C via a pertussis-toxin-sensitive guanine-nucleotide-binding regulatory protein(s) [G-protein(s)]. We have developed methods for the assessment of formyl-peptide-receptor-stimulated binding of radiolabeled guanosine 5'-[gamma-thio]triphosphate ([S-35]GTP[S]) to native HL-60 membranes. Agonist stimulation of [S-35]GTP[S] association with the membrane was minimal (less-than-or-equal-to 20%) when GTP[S] was the sole nucleotide present in the incubation medium. In contrast, receptor activation led to a marked (up to sixfold) stimulation of [S-35]GTP[S] binding when GDP or GTP were present in high (> 100-fold) excess of [S-35]GTP[S]. The increase in [S-35]GTP[S] binding caused by the chemotactic agonist was strictly dependent on the presence of Mg2+ and was significantly increased by Na+. Agonist-independent binding of [S-35]GTP[S] and the increase due to the chemotactic agonist were markedly attenuated by both pertussis and cholera toxin. Comparison of the number of chemotactic-peptide-sensitive [S-35]GTP[S]-binding sites to the number of chemotactic peptide receptors present in HL-60 membranes provided direct evidence that a single formyl-peptide receptor is capable of catalyzing the binding of [S-35]GTP[S] to, and thus the activation of, multiple (up to 20) G-proteins in native plasma membranes.
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页码:725 / 732
页数:8
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