Molecular cloning of porcine alveolar macrophage-derived neutrophil chemotactic factors I and II; identification of porcine IL-8 and another intercrine-alpha protein.
Alveolar macrophages (AM) mediate lung inflammation by producing lipid and peptide molecules that attract neutrophils (PMN) to the lung. Recently we described two porcine proteins called alveolar macrophage-derived chemotactic factors, AMCF-I and -II, that are potent, efficacious, and specific PMN chemoattractants both in vitro and in vivo. We report here the cloning of the full-length cDNAs which code for each protein. Porcine AM were stimulated for 4 h in vitro with Escherichia coli endotoxin (LPS), and a cDNA library was created from poly(A)(+)-selected mRNA. Specific oligonucleotide probes for AMCF-I and AMCF-II were amplified from the porcine AM cDNA library by the polymerase chain reaction using degenerate oligonucleotide primer pairs derived from the N-terminal amino acid sequences of the proteins. These probes were used to isolate 2 full-length cDNAs of 1466 (AMCF-I) and 1515 (AMCF-II) base pairs. Both cDNAs code for proteins with four cysteine residues containing the C-X-C sequence characteristic of the intercrine-alpha family of neutrophil chemoattractants. AMCF-I shares 74% identity with human IL-8 and 84% identity with rabbit IL-8, and likely represents the porcine homologue of IL-8. By contrast, AMCF-II has no obvious human homologue. AMCF-II shares 53% identity with human neutrophil activating peptide 2. Its shared identity with the GRO-related proteins is as high as 61% (rat CINC/GRO), and its shared identity with the 78 amino acid epithelial cell-derived neutrophil activator (ENA-78) is 67%. AMCF-II may represent a new member of the intercrine-alpha family of neutrophil chemoattractants.(ABSTRACT TRUNCATED AT 250 WORDS)
Goodman
R B
RB
Medical Research Service, Seattle Veterans Affairs Medical Center, WA 98108.
Foster
D C
DC
Mathewes
S L
SL
Osborn
S G
SG
Kuijper
J L
JL
Forstrom
J W
JW
Martin
T R
TR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002630:Chemotactic Factors / Q000737:chemistry* / Q000235:genetics*; D003001:Cloning, Molecular; D004247:DNA; D016209:Interleukin-8 / Q000737:chemistry*; D016676:Macrophages, Alveolar / Q000737:chemistry*; D008969:Molecular Sequence Data; D009504:Neutrophils; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013552:Swine
D002630:Chemotactic Factors; D016209:Interleukin-8; D012333:RNA, Messenger; C068711:alveolar macrophage-chemotactic factor-II; D004247:DNA