A chicken homolog of mammalian interleukin-1 beta: cDNA cloning and purification of active recombinant protein.
Upon induction with lipopolysaccharide (LPS) the chicken macrophage cell line HD-11 secretes an activity that stimulates the synthesis of a CXC chemokine in the chicken fibroblast cell line CEC-32. We used a cDNA expression cloning strategy in COS cells to characterize this activity. The isolated cDNA clone codes for a polypeptide of 267 amino acids which lacks a hydrophobic N-terminal domain that could serve as secretory signal. Sequence homology and structural features indicate that this protein is the chicken homolog of mammalian interleukin-1 beta (ChIL-1 beta). Northern blot analysis showed that ChIL-1 beta RNA is quickly induced in blood monocyte-derived macrophages reaching maximal levels within one hour after onset of LPS treatment. To test for biological activity of putative mature ChIL-1 beta, a cDNA fragment comprising amino acids 106 to 267 of the open reading frame was expressed in Escherichia coli so that the resulting polypeptide carried a histidine tag at its N-terminus for easy purification by nickel chelate affinity chromatography. Purified His-ChIL-1 beta potently induced CXC chemokine RNA synthesis in CEC-32 cells. When injected intravenously into adult chickens, it quickly induced a transient increase in serum corticosterone levels.
Weining
K C
KC
Abteilung Virologie, University of Freiburg, Germany.
Sick
C
C
Kaspers
B
B
Staeheli
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002460:Cell Line; D002474:Cell-Free System; D019743:Chemokines, CXC / Q000096:biosynthesis; D002642:Chick Embryo; D002645:Chickens; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli; D006639:Histidine / Q000378:metabolism; D006801:Humans; D007375:Interleukin-1 / Q000235:genetics* / Q000302:isolation & purification* / Q000502:physiology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic