Characterization of a low molecular weight isoform of IL-1 receptor antagonist.
IL-1R antagonist (IL-1Ra) exists in two well-characterized forms, 17-kDa secretory IL-1Ra (sIL-1Ra) and 18-kDa intracellular IL-1Ra (icIL-1Ra), that arise by alternative transcription of the same IL-1Ra gene. A third, lower molecular mass form (approximately 16 kDa) was detected by immunoblot within lysates of a variety of cells, including human monocytes and myelomonocytic cell lines. The 16-kDa isoform was designated icIL-1RaII, and the previously established 18-kDa form was designated icIL-1RaI. Intracellular IL-1RaII bound type I IL-1R up to fivefold less avidly than did sIL-1Ra and icIL-1RaI. Microsequencing of cyanogen bromide fragments of purified icIL-1RaII provided evidence consistent with initiation of protein translation at the second start site in either IL-1Ra mRNA. The results of site-directed mutation experiments established that icIL-1RaII could be derived by alternative translation initiation. In vitro transcription and translation of intact sIL-1Ra cDNA in rabbit reticulocyte lysates led to both pro-sIL-1Ra and icIL-1RaII proteins, whereas transcription and translation of icIL-1RaI cDNA produced both icIL-1RaI and icIL-1RaII proteins. Mutation of the first 5' ATG in sIL-1Ra cDNA led to translation of only icIL-1RaII, while only sIL-1Ra was observed after mutation of the second ATG. These results indicate that icIL-1RaII is a third member of the IL-1Ra family and is a 16-kDa, 143-amino acid intracellular protein derived by alternative translation initiation from either sIL-1Ra mRNA or icIL-1Ra mRNA. The role in biology of either intracellular form of IL-1Ra remains unknown.
Malyak
M
M
Department of Medicine, University of Colorado Health Sciences Center, Denver 80262, USA.
Guthridge
J M
JM
Hance
K R
KR
Dower
S K
SK
Freed
J H
JH
Arend
W P
WP
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein; D007536:Isomerism; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding / Q000276:immunology; D014176:Protein Biosynthesis; D011817:Rabbits; D017472:Receptors, Interleukin-1 / Q000037:antagonists & inhibitors*; D012156:Reticulocytes; D017421:Sequence Analysis; D012795:Sialoglycoproteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
C507763:IL1RN protein, human; C507764:Il1rn protein, mouse; D053590:Interleukin 1 Receptor Antagonist Protein; D017472:Receptors, Interleukin-1; D012795:Sialoglycoproteins