Functional high efficiency expression of cloned leucocyte chemoattractant receptor cDNAs

FEBS Letters

Didsbury, Jolin R.¹  Gerard, Norma²  Gerard, Craig²  Tomhave, Eric¹  Uhing, Ronald J.³  Snyderman, Ralph¹ 

[1] Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA;Ina Sue Perlmutter Research Laboratory, Children's Hospital, Pulmonary Division, Beth Israel Hospital, Department of Pediatrics, Harvard Medical School, Boston, MA 02115, USA;Department of Pathology, Duke University Medical Center, Durham, NC 27710, USA

关键词: Formylpeptide;    Chemoattractant receptor;    Gene expression;    C5a;   

DOI  :  10.1016/0014-5793(92)80555-U

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Human kidney 293 TSA cells were transfected by a calcium phosphate method with human formylpeptide and C5a receptor cDNAs with high efficiency. Formylpeptide receptor positive transfectants expressed a total of 968,000 ± 34,000 receptors per cell with two affinity states (K _ds of ca. 0.43 nM and 39 nM), which in the presence of 100μM GTPγS decreased by ca. 4-fold the number of high-affinity sites. The ligand binding pharmacology of cloned and expressed formylpeptide receptors were indistinguishable from endogenous receptors on human neutrophils. Expressed formylpeptide and C5a receptors were functionally active in mobilizing intracellular calcium via a pertussis toxin sensitive mechanism with an ED₅₀ for formylpeptide of ca. 0.5–1.0 nM. This expression system, in which receptor expression can be monitored by flow cytometric methods and in which intracellular calcium responses are measurable, unlike in the more popular COS-7 cell expression system, will provide a useful basis for the analysis of chemoattractant receptor structure-function relationships.

【 授权许可】

Unknown