Molecular and biochemical characterization of a novel gamma-interferon-inducible protein.
A cDNA clone has been isolated from mRNA derived from the monocytic cell line U937, which detects an mRNA that is present and inducible by gamma-interferon (IFN gamma) in cells of the hematopoietic lineage and absent but inducible in non-hematopoietic cells. Following recombinant IFN gamma (rIFN gamma) treatment, mRNA accumulation can be detected within 30 min, plateaus at 24 h, and remains elevated for several days. This mRNA accumulation is due, at least in part, to increased transcription and is not inhibited by cycloheximide, although cycloheximide alone induces both mRNA accumulation and transcription of this gene. rIFN gamma is a more potent inducing agent than either rIFN alpha or rIFN beta. Polyclonal monospecific antiserum raised to the longest open reading frame expressed in Escherichia coli immunoprecipitates multiple polypeptides that are inducible by rIFN gamma in human monocytes, fibroblasts, endothelial cells, and keratinocytes. In pulse-chase experiments analyzed under reducing conditions, a 30-kDa polypeptide (referred to as IP-30) is either secreted or converted intracellularly into a 25-kDa protein; when analyzed under nonreducing conditions, the two intracellular forms have apparent molecular masses of 25 and 20 kDa, and the extracellular protein is found in two forms of apparent molecular mass 50 and 25 kDa. These data suggest that the intracellular form contains intrachain disulfide bonds, and the extracellular form is involved in both intrachain and interchain disulfide bonding. Indirect immunofluorescence microscopy reveals a punctate fluorescence pattern in monocytes consistent with a vesicular subcellular location. These data are consistent with IP-30 being a novel IFN gamma-inducible protein which may be lysosomal in location.
D001483:Base Sequence; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D003593:Cytoplasm / Q000032:analysis; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004730:Endothelium, Vascular / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D007163:Immunosorbent Techniques; D007371:Interferon-gamma / Q000494:pharmacology*; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000378:metabolism / Q000648:ultrastructure; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014407:Tumor Cells, Cultured
D011506:Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D007371:Interferon-gamma; D004247:DNA; D003513:Cycloheximide