Stable expression of a secreted form of the mouse IFN-gamma receptor by rat cells.
The biologic effects of IFN-gamma are mediated through a receptor that is expressed in relatively low abundance on normal mammalian cells. As a consequence, investigations of the physicochemical and ligand-binding properties of the purified receptor have been limited. The work reported here characterizes a secreted form of the receptor for mouse IFN-gamma, made by deletion of the nucleotides that code for the anchoring domain from a cDNA that encodes the receptor binding protein and its related signal peptide. When transfected into rat XC cells, this construct produced up to approximately 1 mg/liter of a secreted protein that had the characteristics of the native receptor. Both the secreted protein and its mRNA were of sizes that were consistent with loss of the transmembrane region. The protein was detectable by a mAb that is specific for an epitope that is found in the ligand binding site of the receptor for mouse IFN-gamma, as well as by a goat polyclonal IgG that is monospecific for the mouse IFN-gamma R. Supernates that contained the secreted protein blocked binding of IFN-gamma to mouse IFN-gamma R and inhibited in a dose-dependent manner the IFN-gamma-mediated priming of mouse bone marrow culture-derived macrophages for tumor cell killing. Availability of relatively large amounts of a secreted protein that retains ligand-binding activity should facilitate purification and basic studies of the receptor binding protein and could provide new approaches to the treatment/prevention of diseases that arise due to inappropriate response of cells to IFN-gamma. In addition, because this secreted receptor, unlike others, consists of both the extracellular and intracellular domains, it is likely that it will be useful in determining how the cytoplasmic portion of the receptor is involved in receptor function.
Fernando
L P
LP
Department of Microbiology/Molecular Genetics/Immunology, University of Kansas Medical Center, Kansas City 66103.
LeClaire
R D
RD
Obici
S
S
Zavodny
P J
PJ
Russell
S W
SW
Pace
J L
JL
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015152:Blotting, Northern; D003001:Cloning, Molecular; D015870:Gene Expression; D007371:Interferon-gamma / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D011971:Receptors, Immunologic / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017471:Receptors, Interferon; D013329:Structure-Activity Relationship; D014162:Transfection
D012333:RNA, Messenger; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D007371:Interferon-gamma