Neumann D, Kollewe C, Martin M U, Boraschi D
Dompé Research Center, L'Aquila, Italy; andPharmakologie, Medizinische Hochschule Hannover, Hannover, Germany.
J Immunol. 2000 Sep 15;165(6):3350-7. doi: 10.4049/jimmunol.165.6.3350.
IL-1 signaling is mediated by the type I IL-1R (IL-1RI). The nonsignaling type II receptor has a regulatory function, since it reduces IL-1 effects by scavenging free IL-1 molecules. This regulatory function has been demonstrated only for the soluble form, released from the membrane receptor by action of specific proteases, but is still ill-defined for the membrane receptor itself. To assess the function of membrane IL-1RII, a modified IL-1RII cDNA was constructed, in which the cleavable domain was replaced with the corresponding uncleavable sequence of the epidermal growth factor receptor. The human keratinocyte line HaCaT, which does not express wild-type IL-1RII (wtIL-1RII), was stably transfected with this modified cDNA (unconventionally cleavable IL-1RII (uIL-1RII)). Cells transfected with uIL-1RII expressed the membrane form of IL-1RII, but were unable to produce the 60-kDa soluble receptor. Upon analysis of IL-1 responsiveness, parental HaCaT and vector-transfected cells (E27), expressing IL-1RI and the accessory chain IL-1R accessory protein, were responsive to IL-1. Conversely, cells overexpressing wtIL-1RII (811) or uIL-1RII (9D4) showed comparable reduction in responsiveness to both IL-1alpha (bound by membrane and soluble receptors) and IL-1beta (recognized by the membrane receptor only), suggesting that the membrane form of the IL-1RII is mainly responsible for IL-1 inhibition. In contrast with wtIL-1RII, uIL-1RII did not interact with IL-1R accessory protein. Thus, the membrane form of IL-1RII possesses strong IL-1-inhibitory activity, independent of sequestration of the accessory protein and circumscribed to its ligand sink function.
白细胞介素-1（IL-1）信号传导由I型IL-1受体（IL-1RI）介导。无信号传导功能的II型受体具有调节作用，因为它可通过清除游离的IL-1分子来降低IL-1的效应。这种调节功能仅在特定蛋白酶作用下从膜受体释放的可溶性形式中得到证实，而对于膜受体本身仍未明确界定。为了评估膜型IL-1RII的功能，构建了一种修饰的IL-1RII cDNA，其中可裂解结构域被表皮生长因子受体的相应不可裂解序列所取代。不表达野生型IL-1RII（wtIL-1RII）的人角质形成细胞系HaCaT用这种修饰的cDNA（非常规可裂解的IL-1RII（uIL-1RII））进行稳定转染。用uIL-1RII转染的细胞表达膜型IL-1RII，但无法产生60 kDa的可溶性受体。在分析IL-1反应性时，表达IL-1RI和辅助链IL-1R辅助蛋白的亲本HaCaT细胞和载体转染细胞（E27）对IL-1有反应。相反，过表达wtIL-1RII（811）或uIL-1RII（9D4）的细胞对IL-1α（被膜受体和可溶性受体结合）和IL-1β（仅被膜受体识别）的反应性均有类似程度的降低，这表明IL-1RII的膜形式主要负责IL-1的抑制作用。与wtIL-1RII不同，uIL-1RII不与IL-1R辅助蛋白相互作用。因此，IL-1RII的膜形式具有强大的IL-1抑制活性，独立于辅助蛋白的隔离作用，且局限于其配体汇集功能。
