Abstract
The cytokine interleukin 1 alpha (IL-1 alpha) is a critical mediator of the immune and inflammatory responses. A unique determinant of its activity as compared with IL-1 beta may be its association with the plasma membrane. While the biologic activity of "membrane IL-1" has been extensively reported, the mechanism of membrane binding remains unclear. We report that the N terminus of the 31-kDa IL-1 alpha precursor is myristoylated on specific internal lysine residues. Immunoprecipitation of [3H]myristic acid-radiolabeled human monocyte lysates with IgG antibodies to the 31-kDa IL-1 alpha precursor recovered a protein with the physicochemical properties of the IL-1 alpha N-terminal propiece (16 kDa, pI 4.45). Glycyl N-myristoylation of this protein is precluded by the absence of a glycine residue at position 2, suggesting that the propiece is myristoylated on epsilon-amino groups of lysine. To determine which lysine(s) are acylated, a series of synthetic peptides containing all lysines found in the IL-1 alpha N-terminal propiece were used in an in vitro myristoylation assay containing peptide, myristoyl-CoA, and monocyte lysate as enzyme source. Analysis of the reaction products by reverse-phase HPLC and gas-phase sequencing demonstrated the specific myristoylation of Lys-82 and Lys-83, yielding predominantly monoacylated product. A conserved sequence in the IL-1 beta propiece was myristoylated with at least 8-fold less efficiency. Acylation of the IL-1 alpha precursor by a previously unrecognized lysyl epsilon-amino N-myristoyl-transferase activity may facilitate its specific membrane targeting.
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- Aderem A. A., Albert K. A., Keum M. M., Wang J. K., Greengard P., Cohn Z. A. Stimulus-dependent myristoylation of a major substrate for protein kinase C. Nature. 1988 Mar 24;332(6162):362–364. doi: 10.1038/332362a0. [DOI] [PubMed] [Google Scholar]
- Beuscher H. U., Fallon R. J., Colten H. R. Macrophage membrane interleukin 1 regulates the expression of acute phase proteins in human hepatoma Hep 3B cells. J Immunol. 1987 Sep 15;139(6):1896–1901. [PubMed] [Google Scholar]
- Beuscher H. U., Nickells M. W., Colten H. R. The precursor of interleukin-1 alpha is phosphorylated at residue serine 90. J Biol Chem. 1988 Mar 15;263(8):4023–4028. [PubMed] [Google Scholar]
- Bursten S. L., Locksley R. M., Ryan J. L., Lovett D. H. Acylation of monocyte and glomerular mesangial cell proteins. Myristyl acylation of the interleukin 1 precursors. J Clin Invest. 1988 Nov;82(5):1479–1488. doi: 10.1172/JCI113755. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Buss J. E., Solski P. A., Schaeffer J. P., MacDonald M. J., Der C. J. Activation of the cellular proto-oncogene product p21Ras by addition of a myristylation signal. Science. 1989 Mar 24;243(4898):1600–1603. doi: 10.1126/science.2648572. [DOI] [PubMed] [Google Scholar]
- Carruth L. M., Demczuk S., Mizel S. B. Involvement of a calpain-like protease in the processing of the murine interleukin 1 alpha precursor. J Biol Chem. 1991 Jul 5;266(19):12162–12167. [PubMed] [Google Scholar]
- Cerretti D. P., Kozlosky C. J., Mosley B., Nelson N., Van Ness K., Greenstreet T. A., March C. J., Kronheim S. R., Druck T., Cannizzaro L. A. Molecular cloning of the interleukin-1 beta converting enzyme. Science. 1992 Apr 3;256(5053):97–100. doi: 10.1126/science.1373520. [DOI] [PubMed] [Google Scholar]
- Cho W., Tomasselli A. G., Heinrikson R. L., Kézdy F. J. The chemical basis for interfacial activation of monomeric phospholipases A2. Autocatalytic derivatization of the enzyme by acyl transfer from substrate. J Biol Chem. 1988 Aug 15;263(23):11237–11241. [PubMed] [Google Scholar]
- Conlon P. J., Grabstein K. H., Alpert A., Prickett K. S., Hopp T. P., Gillis S. Localization of human mononuclear cell interleukin 1. J Immunol. 1987 Jul 1;139(1):98–102. [PubMed] [Google Scholar]
- Eugui E. M., Almquist S. J. Antibodies against membrane interleukin 1 alpha activate accessory cells to stimulate proliferation of T lymphocytes. Proc Natl Acad Sci U S A. 1990 Feb;87(4):1305–1309. doi: 10.1073/pnas.87.4.1305. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fuhlbrigge R. C., Fine S. M., Unanue E. R., Chaplin D. D. Expression of membrane interleukin 1 by fibroblasts transfected with murine pro-interleukin 1 alpha cDNA. Proc Natl Acad Sci U S A. 1988 Aug;85(15):5649–5653. doi: 10.1073/pnas.85.15.5649. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goldsmith M. R., Rattner E. C., Koehler M. M., Balikov S. R., Bock S. C. Two-dimensional electrophoresis of small-molecular-weight proteins. Anal Biochem. 1979 Oct 15;99(1):33–40. doi: 10.1016/0003-2697(79)90041-1. [DOI] [PubMed] [Google Scholar]
- Gordon J. I. Protein N-myristoylation: simple questions, unexpected answers. Clin Res. 1990 Oct;38(3):517–528. [PubMed] [Google Scholar]
- Graff J. M., Gordon J. I., Blackshear P. J. Myristoylated and nonmyristoylated forms of a protein are phosphorylated by protein kinase C. Science. 1989 Oct 27;246(4929):503–506. doi: 10.1126/science.2814478. [DOI] [PubMed] [Google Scholar]
- Hedo J. A., Collier E., Watkinson A. Myristyl and palmityl acylation of the insulin receptor. J Biol Chem. 1987 Jan 25;262(3):954–957. [PubMed] [Google Scholar]
- Hogquist K. A., Unanue E. R., Chaplin D. D. Release of IL-1 from mononuclear phagocytes. J Immunol. 1991 Oct 1;147(7):2181–2186. [PubMed] [Google Scholar]
- Jacobs E., Gheysen D., Thines D., Francotte M., de Wilde M. The HIV-1 Gag precursor Pr55gag synthesized in yeast is myristoylated and targeted to the plasma membrane. Gene. 1989 Jun 30;79(1):71–81. doi: 10.1016/0378-1119(89)90093-0. [DOI] [PubMed] [Google Scholar]
- Jones T. L., Simonds W. F., Merendino J. J., Jr, Brann M. R., Spiegel A. M. Myristoylation of an inhibitory GTP-binding protein alpha subunit is essential for its membrane attachment. Proc Natl Acad Sci U S A. 1990 Jan;87(2):568–572. doi: 10.1073/pnas.87.2.568. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kamps M. P., Buss J. E., Sefton B. M. Mutation of NH2-terminal glycine of p60src prevents both myristoylation and morphological transformation. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4625–4628. doi: 10.1073/pnas.82.14.4625. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kobayashi Y., Appella E., Yamada M., Copeland T. D., Oppenheim J. J., Matsushima K. Phosphorylation of intracellular precursors of human IL-1. J Immunol. 1988 Apr 1;140(7):2279–2287. [PubMed] [Google Scholar]
- Kobayashi Y., Oppenheim J. J., Matsushima K. Calcium-dependent binding of phosphorylated human pre interleukin 1 alpha to phospholipids. J Biochem. 1990 May;107(5):666–670. doi: 10.1093/oxfordjournals.jbchem.a123105. [DOI] [PubMed] [Google Scholar]
- Kobayashi Y., Yamamoto K., Saido T., Kawasaki H., Oppenheim J. J., Matsushima K. Identification of calcium-activated neutral protease as a processing enzyme of human interleukin 1 alpha. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5548–5552. doi: 10.1073/pnas.87.14.5548. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kurt-Jones E. A., Beller D. I., Mizel S. B., Unanue E. R. Identification of a membrane-associated interleukin 1 in macrophages. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1204–1208. doi: 10.1073/pnas.82.4.1204. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kurt-Jones E. A., Fiers W., Pober J. S. Membrane interleukin 1 induction on human endothelial cells and dermal fibroblasts. J Immunol. 1987 Oct 1;139(7):2317–2324. [PubMed] [Google Scholar]
- Minnich-Carruth L. L., Suttles J., Mizel S. B. Evidence against the existence of a membrane form of murine IL-1 alpha. J Immunol. 1989 Jan 15;142(2):526–530. [PubMed] [Google Scholar]
- Okubo A., Sone S., Tanaka M., Ogura T. Membrane-associated interleukin 1 alpha as a mediator of tumor cell killing by human blood monocytes fixed with paraformaldehyde. Cancer Res. 1989 Jan 15;49(2):265–270. [PubMed] [Google Scholar]
- Pillai S., Baltimore D. Myristoylation and the post-translational acquisition of hydrophobicity by the membrane immunoglobulin heavy-chain polypeptide in B lymphocytes. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7654–7658. doi: 10.1073/pnas.84.21.7654. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rein A., McClure M. R., Rice N. R., Luftig R. B., Schultz A. M. Myristylation site in Pr65gag is essential for virus particle formation by Moloney murine leukemia virus. Proc Natl Acad Sci U S A. 1986 Oct;83(19):7246–7250. doi: 10.1073/pnas.83.19.7246. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Resh M. D., Ling H. P. Identification of a 32K plasma membrane protein that binds to the myristylated amino-terminal sequence of p60v-src. Nature. 1990 Jul 5;346(6279):84–86. doi: 10.1038/346084a0. [DOI] [PubMed] [Google Scholar]
- Stevenson F. T., Bursten S. L., Locksley R. M., Lovett D. H. Myristyl acylation of the tumor necrosis factor alpha precursor on specific lysine residues. J Exp Med. 1992 Oct 1;176(4):1053–1062. doi: 10.1084/jem.176.4.1053. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stevenson F. T., Torrano F., Locksley R. M., Lovett D. H. Interleukin 1: the patterns of translation and intracellular distribution support alternative secretory mechanisms. J Cell Physiol. 1992 Aug;152(2):223–231. doi: 10.1002/jcp.1041520202. [DOI] [PubMed] [Google Scholar]
- Suttles J., Giri J. G., Mizel S. B. IL-1 secretion by macrophages. Enhancement of IL-1 secretion and processing by calcium ionophores. J Immunol. 1990 Jan 1;144(1):175–182. [PubMed] [Google Scholar]
- Thornberry N. A., Bull H. G., Calaycay J. R., Chapman K. T., Howard A. D., Kostura M. J., Miller D. K., Molineaux S. M., Weidner J. R., Aunins J. A novel heterodimeric cysteine protease is required for interleukin-1 beta processing in monocytes. Nature. 1992 Apr 30;356(6372):768–774. doi: 10.1038/356768a0. [DOI] [PubMed] [Google Scholar]
- Tomasselli A. G., Hui J., Fisher J., Zürcher-Neely H., Reardon I. M., Oriaku E., Kézdy F. J., Heinrikson R. L. Dimerization and activation of porcine pancreatic phospholipase A2 via substrate level acylation of lysine 56. J Biol Chem. 1989 Jun 15;264(17):10041–10047. [PubMed] [Google Scholar]
- Towler D. A., Eubanks S. R., Towery D. S., Adams S. P., Glaser L. Amino-terminal processing of proteins by N-myristoylation. Substrate specificity of N-myristoyl transferase. J Biol Chem. 1987 Jan 25;262(3):1030–1036. [PubMed] [Google Scholar]
- Towler D., Glaser L. Protein fatty acid acylation: enzymatic synthesis of an N-myristoylglycyl peptide. Proc Natl Acad Sci U S A. 1986 May;83(9):2812–2816. doi: 10.1073/pnas.83.9.2812. [DOI] [PMC free article] [PubMed] [Google Scholar]