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Characterisation of the RANTES/MIP‐1α receptor (CC CKR‐1) stably transfected in HEK 293 cells and the recombinant ligands
Author(s) -
Proudfoot Amanda E.I.,
Power Christine A.,
Hoogewerf Arlene,
Montjovent Marc-Olivier,
Borlat Frederic,
Wells Timothy N.C.
Publication year - 1995
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(95)01235-x
Subject(s) - hek 293 cells , chemotaxis , transfection , recombinant dna , chemokine , cell culture , chemokine receptor , receptor , microbiology and biotechnology , thp1 cell line , chemistry , biology , biochemistry , gene , genetics
The CC chemokines RANTES and MIP‐1α are known to activate certain leucocytes and leucocytic cell lines. We have produced and fully characterised the recombinant proteins expressed in E. coli . They induce chemotaxis of the pro‐monocytic cell line, THP‐1 and T cells. THP‐1 cells express three of the known CC chemokine receptors. In order to study the activation of a single receptor, we have expressed the shared receptor (CC CKR‐1) for RANTES and MIP‐1α stably in the HEK 293 cell line. We have examined the effects of RANTES and MIP‐1α on the CC CKR‐1 transfectants by equilibrium binding studies and in a chemotaxis assay. RANTES competes for [ 125 I]RANTES with an IC 50 of 0.6 ± 0.23 nM, whereas MIP‐1α competes for its radiolabelled counterpart with an IC 50 of 10 ± 1.6 nM in the transfectants. These affinities are the same as those measured on the THP‐1 cell line. The stably transfected HEK 293 cells respond to both these chemokines in the chemotaxis assay with the same EC 50 values as those measured for THP‐1 cells. This indicates that this cellular response can be mediated through the CC CKR‐1 receptor.