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Interleukin-1-mediated Stabilization of Mouse KC mRNA Depends on Sequences in both 5′- and 3′-Untranslated Regions

2000; Elsevier BV; Volume: 275; Issue: 17 Linguagem: Inglês

10.1074/jbc.275.17.12987

ISSN

1083-351X

Autores

Julie Tebo, Shyamasree Datta, Raj Kishore, Mikhail Kolosov, Jennifer Major, Yoshihiro Ohmori, Thomas A. Hamilton,

Tópico(s)

Immune Cell Function and Interaction

Resumo

mRNA transcribed from the mouse KC chemokine gene accumulated to significantly higher levels in multiple cell types after treatment with interleukin 1α (IL-1α) as compared with tumor necrosis factor-α (TNFα). Although TNFα and IL-1α both signaled the activation of nuclear factor κB and enhanced transcription of the KC gene with equal potency, only IL-1α treatment resulted in stabilization of KC mRNA. Nucleotide sequences that confer sensitivity for IL-1α-mediated mRNA stabilization were identified within the 5′- and 3′-untranslated regions (UTRs) of KC mRNA using transient transfection of chimeric plasmids containing specific portions of KC mRNA linked to the chloramphenicol acetyltransferase (CAT) gene. When plasmids containing either the 3′- or 5′-UTR of KC mRNA were used, the half-life of CAT mRNA was unaltered either in untreated or IL-1α-stimulated cells. In contrast, CAT mRNA transcribed from plasmids that contained both the 5′- and 3′-UTRs of the KC mRNA decayed more rapidly than control CAT mRNA, and this enhanced decay was prevented in cells treated with IL-1α. A cluster of four overlapping AUUUA motifs within the 3′-UTR was required, whereas the 5′-UTR region exhibited orientation dependence. These findings indicate that cooperative function of the two nucleotide sequences involves a distinct signaling pathway used by IL-1α but not TNFα. mRNA transcribed from the mouse KC chemokine gene accumulated to significantly higher levels in multiple cell types after treatment with interleukin 1α (IL-1α) as compared with tumor necrosis factor-α (TNFα). Although TNFα and IL-1α both signaled the activation of nuclear factor κB and enhanced transcription of the KC gene with equal potency, only IL-1α treatment resulted in stabilization of KC mRNA. Nucleotide sequences that confer sensitivity for IL-1α-mediated mRNA stabilization were identified within the 5′- and 3′-untranslated regions (UTRs) of KC mRNA using transient transfection of chimeric plasmids containing specific portions of KC mRNA linked to the chloramphenicol acetyltransferase (CAT) gene. When plasmids containing either the 3′- or 5′-UTR of KC mRNA were used, the half-life of CAT mRNA was unaltered either in untreated or IL-1α-stimulated cells. In contrast, CAT mRNA transcribed from plasmids that contained both the 5′- and 3′-UTRs of the KC mRNA decayed more rapidly than control CAT mRNA, and this enhanced decay was prevented in cells treated with IL-1α. A cluster of four overlapping AUUUA motifs within the 3′-UTR was required, whereas the 5′-UTR region exhibited orientation dependence. These findings indicate that cooperative function of the two nucleotide sequences involves a distinct signaling pathway used by IL-1α but not TNFα. interleukin tumor necrosis factor nuclear factor κB chloramphenicol acetyltransferase actinomycin D ribonuclease protection assay nucleotide(s) AU-rich sequence element The development of an inflammatory response is mediated, in part, by the modulation of gene expression in multiple participating cell types (1.Adams D.O. Hamilton T.A. Gallin J.I. Goldstein I.M. Snyderman R. Inflammation: Basic Principles and Clinical Correlates. Raven Press, Ltd., New York1992: 637-662Google Scholar, 2.Ohmori Y. Hamilton T.A. Pharmacol. Ther. 1994; 63: 235-264Crossref PubMed Scopus (39) Google Scholar, 3.Nathan C.F. Cohn Z.A. Kelly W. Harris E. Ruddy S. Hedge R. Textbook of Rheumatology. W. B. Saunders Co., New York1995: 144-169Google Scholar, 4.Miyajima A. Miyatake S. Schreurs J. De Vries J. Arai N. Yokota T. Arai K. FASEB J. 1988; 2: 2462-2473Crossref PubMed Scopus (135) Google Scholar). The changes in gene expression associated with inflammation are stringently controlled in response to extracellular signals encountered in the microenvironment of the inflammatory site. Among the most potent of such signals are the cytokines interleukin-1α/β (IL-1α/β)1 and tumor necrosis factor-α (TNFα). Although IL-1α and TNFα are structurally unrelated and interact with distinct receptors, they induce a very similar set of functional responses in many cell types (5.Vilcek J. Lee T.H. J. Biol. Chem. 1991; 266: 7313-7316Abstract Full Text PDF PubMed Google Scholar, 6.Le J. Vilcek J. Lab. Invest. 1987; 56: 234-248PubMed Google Scholar, 7.Dinarello C.A. Eur. Cytokine. Netw. 1994; 5: 517-531PubMed Google Scholar, 8.Beutler B. J. Invest. Med. 1995; 43: 227-235PubMed Google Scholar). Indeed, some components of the intracellular signaling pathways that mediate cellular response to TNFα and IL-1α are shared (9.Malinin N.L. Boldin M.P. Kovalenko A.V. Wallach D. Nature. 1997; 385: 540-544Crossref PubMed Scopus (1159) Google Scholar, 10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar, 11.Jobin C. Holt L. Bradham C.A. Streetz K. Brenner D.A. Sartor R.B. J. Immunol. 1999; 162: 4447-4454PubMed Google Scholar, 12.Auron P.E. Cytokine Growth Factor Rev. 1998; 9: 221-237Crossref PubMed Scopus (154) Google Scholar, 13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar). These signaling pathways lead to alterations in gene transcription, in mRNA stability, and in mRNA translation (10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar,13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar, 14.Eder J. Trends Pharmacol. Sci. 1997; 18: 319-322Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar, 15.Holtmann H. Mol. Cell. Biol. 1999; 19: 6742-6753Crossref PubMed Scopus (268) Google Scholar, 16.Saklatvala J. Dean J. Finch A. Biochem. Soc. Symp. 1999; 64: 63-77PubMed Google Scholar).Members of the chemoattractant cytokine or chemokine gene families are recognized as important targets of TNFα/IL-1α during inflammation (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The products of these genes serve to recruit select subsets of inflammatory leukocytes into tissue sites and have significant impact on the magnitude and character of an inflammatory reaction. In the mouse, one of the earliest events in inflammation is the infiltration of neutrophils, a process controlled in part by members of the CXC chemokine family that includes IL-8 and GROα (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). Though there is no direct equivalent of IL-8 in the mouse, the GRO-α-related genes are termed KC and MIP-2 (20.Sherry B. Horii Y. Manogue K.R. Widmer U. Cerami A. Cytokines. 1992; 4: 117-130PubMed Google Scholar, 21.Oquendo P. Albertas J. Wen D. Graycan J.L. Derynk R. Stiles C.D. J. Biol. Chem. 1989; 264: 4133-4137Abstract Full Text PDF PubMed Google Scholar). Expression of the mouse KC gene is stimulated by a number of inflammatory agents including lipopolysaccharide, thrombin, platelet-derived growth factor, TNFα, and IL-1α/β in a number of cell types including macrophages, fibroblasts, epithelial cells, and endothelial cells (22.Hall D.J. Jones S.D. Kaplan D.R. Whitman M. Rollins B.J. Stiles C.D. Mol. Cell. Biol. 1989; 9: 1705-1713Crossref PubMed Scopus (26) Google Scholar, 23.Hall D.J. Brownlee C. Stiles C.D. J. Cell. Physiol. 1989; 141: 154-159Crossref PubMed Scopus (31) Google Scholar, 24.Introna M. Bast Jr., R.C. Tannenbaum C.S. Hamilton T.A. Adams D.O. J. Immunol. 1987; 11: 3891-3896Google Scholar, 25.Shen X.Y. Hamilton T.A. DiCorleto P.E. J. Cell. Physiol. 1989; 140: 44-51Crossref PubMed Scopus (22) Google Scholar, 26.Shen X.Y. Hamilton T.A. DiCorleto P.E. Biochim. Biophys. Acta. 1990; 2: 145-150Crossref Scopus (5) Google Scholar). Although KC mRNA expression requires increased KC gene transcription through the action of NFκB, IL-1α has been shown to enhance the accumulation of KC mRNA by prolonging its half-life in BALB/c 3T3 cells (27.Ohmori Y. Hamilton T.A. Biochem. Biophys. Res. Commun. 1994; 2: 590-596Crossref Scopus (77) Google Scholar, 28.Ohmori Y. Fukumoto S. Hamilton T.A. J. Immunol. 1995; 155: 3593-3600PubMed Google Scholar).In the present study we consider the mechanism(s) by which IL-1α can selectively stabilize KC mRNA. Although TNFα and IL-1α are both competent stimuli for the activation of NFκB and associated changes in gene transcription, only IL-1α stabilizes KC mRNA. Furthermore, both the constitutive instability of KC mRNA and the sensitivity to IL-1α depend upon two separate nucleotide sequences within the mature mRNA. Although a cluster of four overlapping AUUUA pentamers in the 3′-UTR is required for both rapid decay and IL-1α-mediated stabilization, this sequence motif is by itself insufficient and requires the 68 nucleotides composing the 5′-UTR as well.DISCUSSIONChemokines are now believed to be critical players in the development and resolution of inflammatory reactions (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The overlapping target cell specificity and receptor utilization of many chemokines, however, suggest that the control of chemokine expressionin vivo may be an important determinant of their distinct functions (19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The expression of many chemokine genes is known to be controlled at the level of transcription, and the mechanisms involved are well studied (27.Ohmori Y. Hamilton T.A. Biochem. Biophys. Res. Commun. 1994; 2: 590-596Crossref Scopus (77) Google Scholar, 28.Ohmori Y. Fukumoto S. Hamilton T.A. J. Immunol. 1995; 155: 3593-3600PubMed Google Scholar, 42.Ohmori Y. Wyner L. Narumi S. Armstrong D. Stoler M. Hamilton T.A. Am. J. Pathol. 1993; 142: 861-870PubMed Google Scholar, 43.Ohmori Y. Hamilton T.A. J. Biol. Chem. 1993; 268: 6677-6688Abstract Full Text PDF PubMed Google Scholar, 44.Yasumoto K. Okamoto S. Mukaida N. Murakami S. Mai M. Matsushima K. J. Biol. Chem. 1992; 267: 22506-22511Abstract Full Text PDF PubMed Google Scholar, 45.Matsusaka T. Fujikawa K. Nishio Y. Mukaida N. Matsushima K. Kishimoto T. Akira S. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10193-10197Crossref PubMed Scopus (873) Google Scholar). It is likely, however, that post-transcriptional mechanisms are also important. Indeed, prior work from this and other laboratories has demonstrated that stimulus-dependent changes in specific chemokine mRNA stability can be an important and even critical determinant of the level of specific chemokine gene expression (27.Ohmori Y. Hamilton T.A. Biochem. Biophys. Res. Commun. 1994; 2: 590-596Crossref Scopus (77) Google Scholar, 46.Kasama T. Strieter R.M. Lukacs N.W. Burdick M.D. Kunkel S.L. J. Immunol. 1994; 152: 3559-3569PubMed Google Scholar, 47.Kim H.S. Armstrong D. Hamilton T.A. Tebo J.M. J. Leukocyte Biol. 1998; 64: 33-39Crossref PubMed Scopus (50) Google Scholar, 48.Kishore R. Tebo J.M. Kolosov M. Hamilton T.A. J. Immunol. 1999; 162: 2457-2461PubMed Google Scholar). In the present study, we have explored the IL-1α-mediated post-transcriptional regulation of mouse KC gene expression. The results of these experiments support the following conclusions. 1) IL-1α-mediated stabilization of KC mRNA is a general phenomenon that can be detected in fibroblast, epithelial, and endothelial cell types. 2) The IL-1α-initiated intracellular signaling pathway leading to the activation of NFκB is not sufficient for inducing alterations in KC mRNA stability. 3) Nucleotide sequences in the 5′-UTR and 3′-UTR of KC mRNA are both required for destabilizing the message and for conferring sensitivity to IL-1α for mRNA stabilization.IL-1α/β and TNFα are known to exhibit broadly overlapping functional activities that include the ability to modulate proinflammatory gene expression in a wide array of cell types (5.Vilcek J. Lee T.H. J. Biol. Chem. 1991; 266: 7313-7316Abstract Full Text PDF PubMed Google Scholar, 6.Le J. Vilcek J. Lab. Invest. 1987; 56: 234-248PubMed Google Scholar, 7.Dinarello C.A. Eur. Cytokine. Netw. 1994; 5: 517-531PubMed Google Scholar, 8.Beutler B. J. Invest. Med. 1995; 43: 227-235PubMed Google Scholar). Not surprisingly, these agents also share many intracellular signaling components (9.Malinin N.L. Boldin M.P. Kovalenko A.V. Wallach D. Nature. 1997; 385: 540-544Crossref PubMed Scopus (1159) Google Scholar, 10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar, 11.Jobin C. Holt L. Bradham C.A. Streetz K. Brenner D.A. Sartor R.B. J. Immunol. 1999; 162: 4447-4454PubMed Google Scholar, 12.Auron P.E. Cytokine Growth Factor Rev. 1998; 9: 221-237Crossref PubMed Scopus (154) Google Scholar, 13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar). For example, both TNFα and IL-1 are known to stimulate the activation of NFκB, and data presented here demonstrate that, under the experimental conditions employed, both agents were equipotent stimuli for the induced degradation of IκBα, the activation of κB sequence-specific DNA binding activity, and the stimulation of NFκB-dependent gene transcription. Receptor occupancy by either IL-1α or TNFα leads to a series of sequential protein-protein interactions that are distinct for each ligand receptor pair (10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar, 12.Auron P.E. Cytokine Growth Factor Rev. 1998; 9: 221-237Crossref PubMed Scopus (154) Google Scholar, 13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar). Both pathways, however, converge with involvement of TRAF6 and share in common the downstream components NIK, the IκB kinase complex, and finally the release and nuclear translocation of NFκB. The finding that IL-1α but not TNFα can stimulate the stabilization of KC mRNA clearly demonstrates that IL-1α is able to activate signaling events that are not shared with TNFα. In this regard, TNFα and IL-1 have been reported to produce differential effects in some cell types and/or to use nonidentical signaling pathways to achieve the same biologic outcomes (14.Eder J. Trends Pharmacol. Sci. 1997; 18: 319-322Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar, 16.Saklatvala J. Dean J. Finch A. Biochem. Soc. Symp. 1999; 64: 63-77PubMed Google Scholar, 49.Muzio M. Natoli G. Saccani S. Levrero M. Mantovani A. J. Exp. Med. 1998; 187: 2097-2101Crossref PubMed Scopus (525) Google Scholar). At least a portion of this diversity is likely to involve the contribution of the stress-activated/mitogen-activated protein kinase signaling pathways (14.Eder J. Trends Pharmacol. Sci. 1997; 18: 319-322Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar, 16.Saklatvala J. Dean J. Finch A. Biochem. Soc. Symp. 1999; 64: 63-77PubMed Google Scholar, 49.Muzio M. Natoli G. Saccani S. Levrero M. Mantovani A. J. Exp. Med. 1998; 187: 2097-2101Crossref PubMed Scopus (525) Google Scholar), and recent reports link these signaling pathways with regulation of mRNA stability (50.Chen C.Y. Del Gatto-Konczak F. Wu Z. Karin M. Science. 1998; 280: 1945-1949Crossref PubMed Scopus (329) Google Scholar, 51.Ridley S.H. Dean J.L. Sarsfield S.J. Brook M. Clark A.R. Saklatvala J. FEBS Lett. 1998; 439: 75-80Crossref PubMed Scopus (201) Google Scholar).Multiple studies demonstrate that IL-1 can modulate the stability of specific mRNAs, including human GRO-α, IL-8, IL-6, COX2, and IL-11 (52.Stoeckle M.Y. Nucleic Acids Res. 1991; 19: 917-920Crossref PubMed Scopus (76) Google Scholar, 53.Stoeckle M.Y. Nucleic Acids Res. 1992; 20: 1123-1127Crossref PubMed Scopus (50) Google Scholar, 54.Ng S.B. Tan Y.H. Guy G.R. J. Biol. Chem. 1994; 269: 19021-19027Abstract Full Text PDF PubMed Google Scholar, 55.Yang L. Yang Y.C. J. Biol. Chem. 1994; 269: 32732-32739Abstract Full Text PDF PubMed Google Scholar). This activity apparently is mediated through specific regulatory sequences present in the mature mRNAs. As is true for regulatory sequences in genomic DNA that control transcription, there can be multiple elements in a particular mRNA that determine, either independently or cooperatively, the half-life of the molecule (50.Chen C.Y. Del Gatto-Konczak F. Wu Z. Karin M. Science. 1998; 280: 1945-1949Crossref PubMed Scopus (329) Google Scholar, 55.Yang L. Yang Y.C. J. Biol. Chem. 1994; 269: 32732-32739Abstract Full Text PDF PubMed Google Scholar). Indeed regions in both the 5′-UTR and the 3′-UTR of mRNAs can function in cooperative fashion to confer sensitivity to various stimuli (50.Chen C.Y. Del Gatto-Konczak F. Wu Z. Karin M. Science. 1998; 280: 1945-1949Crossref PubMed Scopus (329) Google Scholar, 55.Yang L. Yang Y.C. J. Biol. Chem. 1994; 269: 32732-32739Abstract Full Text PDF PubMed Google Scholar). The present findings confirm these prior observations and provide more precise determination of the nature of the sequence regions that are required. Although both the 68-nucleotide 5′-UTR and a portion of the 3′-UTR containing an ARE cluster are required, neither fragment alone alters the metabolic behavior of the reporter mRNA molecule; only when both sequences are present is the mRNA destabilized and sensitive to IL-1α-mediated stabilization. Whether these sites are recognized independently by distinct proteins or function as part of a larger complex remains to be determined.It is noteworthy that the sequence motif in the KC 3′-UTR that is necessary for destabilization and IL-1α sensitivity contains a classic AU-rich motif composed of four overlapping AUUUA pentamers. Such sequence motifs are well known to be important determinants of the instability of many growth factor and cytokine mRNAs (38.Ross J. Microbiol. Rev. 1995; 3: 423-450Crossref Google Scholar, 39.Chen C.-Y.A. Shyu A.-B. Trends Biochem. Sci. 1995; 20: 465-470Abstract Full Text PDF PubMed Scopus (1666) Google Scholar, 40.Caput D. Beutler B. Hartog K. Thayer R. Brown-Shimer S. Cerami A. Proc. Natl. Acad. Sci. U. S. A. 1986; 83: 1670-1674Crossref PubMed Scopus (1207) Google Scholar, 41.Shaw G. Kamen R. Cell. 1986; 46: 659-667Abstract Full Text PDF PubMed Scopus (3106) Google Scholar). Studies of the mechanisms controlling mRNA stability demonstrate that shortening of the polyadenylated mRNA tail may be the first step in targeting an mRNA for degradation, and this is determined, at least in part, by the nature of the ARE motif (56.Xu N. Chen C.Y. Shyu A.B. Mol. Cell. Biol. 1997; 17: 4611-4621Crossref PubMed Scopus (307) Google Scholar). Whether sensitivity for IL-1-mediated modulation of stability is a property of all AREs or is restricted to a subset is at present not known.Furthermore, there have been many reports identifying RNA-binding proteins that exhibit sequence specificity for AREs (57.Hel Z. Skamene E. Radzioch D. Mol. Cell. Biol. 1996; 16: 5579-5590Crossref PubMed Scopus (46) Google Scholar, 58.Zhang W. Wagner B.J. Ehrenman K. Schaefer A.W. De Maria C.T. Crater D. De Haven K. Long L. Brewer G. Mol. Cell. Biol. 1993; 13: 7652-7665Crossref PubMed Scopus (492) Google Scholar, 59.Myer V.E. Fan X.C. Steitz J.A. EMBO J. 1997; 16: 2130-2139Crossref PubMed Scopus (279) Google Scholar, 60.Antic D. Keene J.D. Am. J. Hum. Genet. 1997; 61: 273-278Abstract Full Text PDF PubMed Scopus (212) Google Scholar, 61.Wilson G.M. Brewer G. Prog. Nucleic Acid Res. Mol. Biol. 1999; 62: 257-291Crossref PubMed Scopus (123) Google Scholar). At least two families of ARE-binding proteins have been well characterized. The Elav family (embryonic lethal abnormal vision) contains at least four members (elr A-D), one of which, termed HuR or elr A, is widely expressed and has been clearly demonstrated to bind AREs and regulate the stability of ARE-containing mRNAs (62.Fan X.C. Steitz J.A. EMBO J. 1998; 17: 3448-3460Crossref PubMed Scopus (743) Google Scholar, 63.Peng S.S. Chen C.Y. Xu N. Shyu A.B. EMBO J. 1998; 17: 3461-3470Crossref PubMed Scopus (652) Google Scholar, 64.Ford L.P. Watson J. Keene J.D. Wilusz J. Genes Dev. 1999; 13: 188-201Crossref PubMed Scopus (219) Google Scholar). A second well characterized RNA-binding protein is termed AUF1 and is a member of the heterogeneous nuclear ribonucleoprotein family (65.Wagner B.J. DeMaria C.T. Sun Y. Wilson G.M. Brewer G. Genomics. 1998; 48: 195-202Crossref PubMed Scopus (233) Google Scholar). The binding of the HuR protein with an ARE containing mRNA is correlated with mRNA stabilization, while interaction with AUF1 appears to destabilize the target mRNA (62.Fan X.C. Steitz J.A. EMBO J. 1998; 17: 3448-3460Crossref PubMed Scopus (743) Google Scholar, 63.Peng S.S. Chen C.Y. Xu N. Shyu A.B. EMBO J. 1998; 17: 3461-3470Crossref PubMed Scopus (652) Google Scholar, 64.Ford L.P. Watson J. Keene J.D. Wilusz J. Genes Dev. 1999; 13: 188-201Crossref PubMed Scopus (219) Google Scholar, 66.De Maria C.T. Brewer G. J. Biol. Chem. 1996; 271: 12179-12184Abstract Full Text Full Text PDF PubMed Scopus (237) Google Scholar, 67.Loflin P. Chen C.Y. Shyu A.B. Genes Dev. 1999; 13: 1884-1897Crossref PubMed Scopus (262) Google Scholar). Which, if either, of these proteins participates in the mechanism of IL-1α-mediated KC mRNA stabilization will be the object of future studies. The development of an inflammatory response is mediated, in part, by the modulation of gene expression in multiple participating cell types (1.Adams D.O. Hamilton T.A. Gallin J.I. Goldstein I.M. Snyderman R. Inflammation: Basic Principles and Clinical Correlates. Raven Press, Ltd., New York1992: 637-662Google Scholar, 2.Ohmori Y. Hamilton T.A. Pharmacol. Ther. 1994; 63: 235-264Crossref PubMed Scopus (39) Google Scholar, 3.Nathan C.F. Cohn Z.A. Kelly W. Harris E. Ruddy S. Hedge R. Textbook of Rheumatology. W. B. Saunders Co., New York1995: 144-169Google Scholar, 4.Miyajima A. Miyatake S. Schreurs J. De Vries J. Arai N. Yokota T. Arai K. FASEB J. 1988; 2: 2462-2473Crossref PubMed Scopus (135) Google Scholar). The changes in gene expression associated with inflammation are stringently controlled in response to extracellular signals encountered in the microenvironment of the inflammatory site. Among the most potent of such signals are the cytokines interleukin-1α/β (IL-1α/β)1 and tumor necrosis factor-α (TNFα). Although IL-1α and TNFα are structurally unrelated and interact with distinct receptors, they induce a very similar set of functional responses in many cell types (5.Vilcek J. Lee T.H. J. Biol. Chem. 1991; 266: 7313-7316Abstract Full Text PDF PubMed Google Scholar, 6.Le J. Vilcek J. Lab. Invest. 1987; 56: 234-248PubMed Google Scholar, 7.Dinarello C.A. Eur. Cytokine. Netw. 1994; 5: 517-531PubMed Google Scholar, 8.Beutler B. J. Invest. Med. 1995; 43: 227-235PubMed Google Scholar). Indeed, some components of the intracellular signaling pathways that mediate cellular response to TNFα and IL-1α are shared (9.Malinin N.L. Boldin M.P. Kovalenko A.V. Wallach D. Nature. 1997; 385: 540-544Crossref PubMed Scopus (1159) Google Scholar, 10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar, 11.Jobin C. Holt L. Bradham C.A. Streetz K. Brenner D.A. Sartor R.B. J. Immunol. 1999; 162: 4447-4454PubMed Google Scholar, 12.Auron P.E. Cytokine Growth Factor Rev. 1998; 9: 221-237Crossref PubMed Scopus (154) Google Scholar, 13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar). These signaling pathways lead to alterations in gene transcription, in mRNA stability, and in mRNA translation (10.Wallach D. Varfolomeev E.E. Malinin N.L. Goltsev Y.V. Kovalenko A.V. Boldin M.P. Annu. Rev. Immunol. 1999; 17: 331-367Crossref PubMed Scopus (1121) Google Scholar,13.O'Neill L.A. Greene C. J. Leukocyte Biol. 1998; 63: 650-657Crossref PubMed Scopus (494) Google Scholar, 14.Eder J. Trends Pharmacol. Sci. 1997; 18: 319-322Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar, 15.Holtmann H. Mol. Cell. Biol. 1999; 19: 6742-6753Crossref PubMed Scopus (268) Google Scholar, 16.Saklatvala J. Dean J. Finch A. Biochem. Soc. Symp. 1999; 64: 63-77PubMed Google Scholar). Members of the chemoattractant cytokine or chemokine gene families are recognized as important targets of TNFα/IL-1α during inflammation (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The products of these genes serve to recruit select subsets of inflammatory leukocytes into tissue sites and have significant impact on the magnitude and character of an inflammatory reaction. In the mouse, one of the earliest events in inflammation is the infiltration of neutrophils, a process controlled in part by members of the CXC chemokine family that includes IL-8 and GROα (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). Though there is no direct equivalent of IL-8 in the mouse, the GRO-α-related genes are termed KC and MIP-2 (20.Sherry B. Horii Y. Manogue K.R. Widmer U. Cerami A. Cytokines. 1992; 4: 117-130PubMed Google Scholar, 21.Oquendo P. Albertas J. Wen D. Graycan J.L. Derynk R. Stiles C.D. J. Biol. Chem. 1989; 264: 4133-4137Abstract Full Text PDF PubMed Google Scholar). Expression of the mouse KC gene is stimulated by a number of inflammatory agents including lipopolysaccharide, thrombin, platelet-derived growth factor, TNFα, and IL-1α/β in a number of cell types including macrophages, fibroblasts, epithelial cells, and endothelial cells (22.Hall D.J. Jones S.D. Kaplan D.R. Whitman M. Rollins B.J. Stiles C.D. Mol. Cell. Biol. 1989; 9: 1705-1713Crossref PubMed Scopus (26) Google Scholar, 23.Hall D.J. Brownlee C. Stiles C.D. J. Cell. Physiol. 1989; 141: 154-159Crossref PubMed Scopus (31) Google Scholar, 24.Introna M. Bast Jr., R.C. Tannenbaum C.S. Hamilton T.A. Adams D.O. J. Immunol. 1987; 11: 3891-3896Google Scholar, 25.Shen X.Y. Hamilton T.A. DiCorleto P.E. J. Cell. Physiol. 1989; 140: 44-51Crossref PubMed Scopus (22) Google Scholar, 26.Shen X.Y. Hamilton T.A. DiCorleto P.E. Biochim. Biophys. Acta. 1990; 2: 145-150Crossref Scopus (5) Google Scholar). Although KC mRNA expression requires increased KC gene transcription through the action of NFκB, IL-1α has been shown to enhance the accumulation of KC mRNA by prolonging its half-life in BALB/c 3T3 cells (27.Ohmori Y. Hamilton T.A. Biochem. Biophys. Res. Commun. 1994; 2: 590-596Crossref Scopus (77) Google Scholar, 28.Ohmori Y. Fukumoto S. Hamilton T.A. J. Immunol. 1995; 155: 3593-3600PubMed Google Scholar). In the present study we consider the mechanism(s) by which IL-1α can selectively stabilize KC mRNA. Although TNFα and IL-1α are both competent stimuli for the activation of NFκB and associated changes in gene transcription, only IL-1α stabilizes KC mRNA. Furthermore, both the constitutive instability of KC mRNA and the sensitivity to IL-1α depend upon two separate nucleotide sequences within the mature mRNA. Although a cluster of four overlapping AUUUA pentamers in the 3′-UTR is required for both rapid decay and IL-1α-mediated stabilization, this sequence motif is by itself insufficient and requires the 68 nucleotides composing the 5′-UTR as well. DISCUSSIONChemokines are now believed to be critical players in the development and resolution of inflammatory reactions (17.Luster A.D. N. Engl. J. Med. 1998; 338: 436-445Crossref PubMed Scopus (3231) Google Scholar, 18.Baggiolini M. Nature. 1998; 392: 565-568Crossref PubMed Scopus (2384) Google Scholar, 19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The overlapping target cell specificity and receptor utilization of many chemokines, however, suggest that the control of chemokine expressionin vivo may be an important determinant of their distinct functions (19.Rollins B.J. Blood. 1997; 90: 909-928Crossref PubMed Google Scholar). The expression of many chemokine genes is known to be controlled at the level of transcription, and the mechanisms involved are well studied (27.Ohmori Y. Hamilton T.A. Biochem. Biophys. Res. Commun. 1994; 2: 590-596Crossref Scopus (77) Google Scholar, 28.Ohmori Y. Fukumoto S. Hamilton T.A. J. Immunol. 1995; 155: 3593-3600PubMed Google Scholar, 42.Ohmori Y. Wyner L. Narumi S. Armstrong D. Stoler M. Hamilton T.A. Am. J. Pathol. 1993; 142: 861-870PubMed Google Scholar, 43.Ohmori Y. Hamilton T.A. J. Biol. Chem. 1993; 268: 6677-6688Abstract Full Text PDF PubMed Google Scholar, 44.Yasumoto K. Okamoto S. Mukaida N. Murakami S. Mai M. Matsushima K. J. Biol. Chem. 1992; 267: 22506-22511Abstract Full Text PDF PubMed Google Scholar, 45.Matsusaka T. Fujikawa K. Nishio Y. Mukaida N. Matsushima K. Kishimoto T. Akira S. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10193-10197Crossref PubMed Scopus (873) Google Scholar). It is likely, however, that post-transcriptional mechanisms are also important.

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