Kostura M J, Tocci M J, Limjuco G, Chin J, Cameron P, Hillman A G, Chartrain N A, Schmidt J A
Department of Biochemical and Molecular Pathology, Merck Sharp & Dohme Research Laboratories, Rahway, NJ 07065.
Proc Natl Acad Sci U S A. 1989 Jul;86(14):5227-31. doi: 10.1073/pnas.86.14.5227.
Interleukin 1 (IL-1) is a lymphokine secreted by monocytes in response to a variety of inflammatory stimuli. IL-1 beta, the predominant form of IL-1 produced by human monocytes, is synthesized as an inactive precursor of 31 kDa and is cleaved at Asp116-Ala117 to yield a 17.5-kDa extracellular form. The exact cellular site of cleavage and mechanism of secretion is at present unknown. We have prepared cell-free postnuclear extracts from freshly isolated human monocytes as well as THP.1 cells, a human monocyte-like cell line, and various blood lymphocytes and fibroblast cell lines. Using pre-IL-1 beta synthesized by in vitro transcription and translation, we have shown that only extracts derived from human monocytes and THP.1 cells were capable of cleaving precursor IL-1 beta to authentic mature IL-1 beta. Subcellular fractionation of the extracts suggested that the processing activity is found in the cytosol of monocytes or monocyte-like cell lines. The cleavage product of this protease is identical to authentic IL-1 beta as shown by mobility on SDS/PAGE and amino acid sequence analysis of the [3H]leucine-labeled product. The cleavage product is also capable of binding to the IL-1 receptor found on fibroblast membranes. Finally, mutation of Asp116----Ala116 rendered the IL-1 beta precursor resistant to cleavage by the processing activity. We conclude that a protease activity found only in monocytes will specifically process IL-1 beta to an active form.
白细胞介素1（IL-1）是单核细胞在受到多种炎症刺激时分泌的一种淋巴因子。IL-1β是人类单核细胞产生的主要IL-1形式，它以31 kDa的无活性前体形式合成，并在Asp116-Ala117处裂解，产生17.5 kDa的细胞外形式。目前尚不清楚裂解的确切细胞位点和分泌机制。我们从新鲜分离的人类单核细胞以及THP.1细胞（一种人类单核细胞样细胞系）、各种血液淋巴细胞和成纤维细胞系中制备了无细胞核后提取物。使用通过体外转录和翻译合成的前体IL-1β，我们发现只有源自人类单核细胞和THP.1细胞的提取物能够将前体IL-1β裂解为真正成熟的IL-1β。提取物的亚细胞分级分离表明，加工活性存在于单核细胞或单核细胞样细胞系的胞质溶胶中。如SDS/PAGE上的迁移率和[3H]亮氨酸标记产物的氨基酸序列分析所示，这种蛋白酶的裂解产物与真正的IL-1β相同。该裂解产物也能够与成纤维细胞膜上发现的IL-1受体结合。最后，将Asp116突变为Ala116使IL-1β前体对加工活性的裂解具有抗性。我们得出结论，仅在单核细胞中发现的一种蛋白酶活性将特异性地将IL-1β加工成活性形式。
