Identification of surrogate agonists for the human FPRL-1 receptor by autocrine selection in yeast.
We describe a procedure for isolating agonists for mammalian G protein-coupled receptors of unknown function. Human formyl peptide receptor like-1 (FPRL-1) receptor, originally identified as an orphan G protein-coupled receptor related to the formyl peptide receptor (FPR1), was expressed in Saccharomyces cells designed to couple receptor activation to histidine prototrophy. Selection for histidine prototrophs among transformants obtained with a plasmid-based library encoding random peptides identified six different agonists, each of whose production yielded autocrine stimulation of the receptor expressed in yeast. A synthetic version of each peptide promoted activation of FPRL-1 expressed in human embryonic kidney (HEK293) cells, and five of the peptides exhibited significant selectivity for activation of FPRL-1 relative to FPR1. One selective peptide was tested and found to mobilize calcium in isolated human neutrophils. This demonstrates that stimulation of FPRL-1 results in neutrophil activation and suggests that the receptor functions as a component of the inflammatory response. This autocrine selection protocol may be a generally applicable method for providing pharmacological tools to evaluate the physiological roles of the growing number of mammalian orphan G protein-coupled receptors.
Klein
C
C
Cadus Pharmaceutical Corporation, Tarrytown, NY 10591-6705, USA. christine.klein@cadus.com
Paul
J I
JI
Sauvé
K
K
Schmidt
M M
MM
Arcangeli
L
L
Ransom
J
J
Trueheart
J
J
Manfredi
J P
JP
Broach
J R
JR
Murphy
A J
AJ
D000595:Amino Acid Sequence; D002460:Cell Line; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000378:metabolism; D009504:Neutrophils / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic / Q000819:agonists* / Q000378:metabolism; D018000:Receptors, Peptide / Q000819:agonists* / Q000378:metabolism; D011994:Recombinant Proteins / Q000819:agonists / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D008024:Ligands; D010455:Peptides; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; D011994:Recombinant Proteins; D009240:N-Formylmethionine Leucyl-Phenylalanine