|[Frontiers in Bioscience 1, d340-357, December 1, 1996]|
MOLECULAR AND CELLULAR BIOLOGY OF INTERLEUKIN-6 AND ITS RECEPTOR|
Evan T. Keller1,2,3, Jon Wanagat1, W.B. Ershler1,3
1Glennan Center for Geriatrics and Gerontology, Departments of 2Pathology and 3Internal Medicine, Eastern Virginia Medical School.
Received 10/21/96; Accepted 11/01/96; On-line //96
IL-6 is involved in a myriad of biologic processes, perhaps explaining its long list of synonyms (B-cell stimulatory factor-2, B cell differentiation factor, T cell-replacing factor, interferon-ß2, 26-kDa protein, hybridoma growth factor, interleukin hybridoma plasmacytoma factor 1, plasmacytoma growth factor, hepatocyte-stimulating factor, macrophage granulocyte-inducing factor 2, cytotoxic T cell differentiation factor, thrombopoietin) (1). Though not an exclusive representation, the several biologic activities of IL-6 are depicted in Figure 1.
Figure 1: Roles of Interleukin-6 in Physiology
Interleukin-6, termed at the time interferon-ß2, was first cloned during an effort to isolate and characterize the viral-induced protein interferon-ß. This strategy included treating cultured human fibroblasts with the double-stranded RNA, poly(I)·poly(C), which mimics viral activity (2). IL-6 is now well recognized for its role in the acute phase inflammatory response which is characterized by production of a variety of hepatic proteins termed acute phase proteins (e.g., C-reactive protein, serum amyloid A, fibrinogen, complement, alpha1-antitrypsin) (reviewed in (3)). In addition to its role in the acute phase response, IL-6 is important for the development of specific immunologic responses. IL-6 induces differentiation of activated, but not resting, B cells (4-6) culminating in production of immunoglobulin (7, 8). Along with B cell differentiation, IL-6 stimulates proliferation of thymic and peripheral T cells (9, 10) and in cooperation with IL-1 (11), induces T cell differentiation to cytolytic-T cells (12, 13) and activates natural killer cells (14). These observations emphasize the importance of IL-6 in both non-specific and specific immune responses.
In addition to its immunologic/inflammatory role, IL-6 appears to play an important role in bone metabolism through induction of osteoclastogenesis and osteoclast activity (15, 16). In rodents, inhibition of IL-6 gene expression is in part responsible for estrogen's ability to inhibit osteoclast activation (17-20). These findings are further supported by the observation that IL-6 gene knockout mice are protected from cancellous bone loss associated with ovariectomy (21).
In addition to the activities described above, IL-6 functions in a wide variety of other systems including the reproductive system by participating in menses (22, 23) and spermatogenesis (24), skin proliferation (25-27), megakaryocytopoiesis (28-30), macrophage differentiation (31-33), and neural cell differentiaion and proliferation (34, 35).
Because of its multidimensional and complex actions, dysregulation of IL-6 results in a myriad of disorders (summarized in Fig. 2) including a variety of neoplastic processes. For example, it may affect cancer progression by its actions on cell adhesion and motility (36), thrombopoiesis (30, 37), tumor specific antigen expression (38) and cancer cell proliferation. Depending on the cell type and the presence or absence of IL-6R, IL-6 can either inhibit (39-41) or stimulate (42) cancer cell proliferation. A great variety of tumor types are stimulated by IL-6, including melanoma (43), renal cell carcinoma (44, 45), prostate carcinoma (46), Kaposi's sarcoma (47), ovarian carcinoma (48), lymphoma and leukemia (49-51), and multiple myeloma (52-59). In many of these tumors, IL-6R have been detected and a direct proliferative signal has been proposed. Yet, when tumor cells are devoid of IL-6R, a tumor inhibiting effect of IL-6 has been demonstrated, presumably because of its immune enhancing properties.
Figure 2: Roles of Interleukin-6 in Pathophysiology
Recently, IL-6 like other cytokines and growth factors (e.g., IL1-alpha, IL1-ß, and TNF-alpha) has been shown to contribute to the bone remodeling process (for review see references (60, 61)). IL-6 exerts its effect on bone by stimulating osteoclast progenitor cell differentiation and osteoclast proliferation as mentioned above. Conditioned media from marrow cultures obtained from patients with Paget's disease (characterized by increased osteoclastogenesis), stimulated osteoclast-like cell formation in normal human marrow cultures and this was reversed by addition of neutralizing antibody to IL-6 (62). IL-6 neutralizing antibody also blocks bone resorption induced by a variety of agents including TNF (18, 63). In addition to increasing osteoclast numbers, IL-6 has been shown to stimulate bone resorption in rat long bones (64) and fetal mouse metacarpi (65) , calvaria (66), and bone resorption pit assays (62, 67). Although it is not clear that IL-6 alone is sufficient to mediate these activities (68), these data demonstrate the importance of IL-6 in enhancing osteoclastic activity thus providing a mechanism for IL-6 promoting osteoporosis.
Although a normal physiologic process, aging is accompanied by a variety of disorders (reviewed in (69)) including, Alzheimer's disease, arteriosclerosis, and thyroiditis. Because IL-6 levels are directly correlated with aging in a variety of species (reviewed in (70)), it may play an important role in the aging process. Intriguingly, dietary restriction, the only experimental intervention that reproducibly prolongs maximum lifespan in mammals (71) can restore to the young phenotype a variety of physiologic parameters, including IL-6 secretion and serum levels.(72, 73). Similarly, DHEA, currently thought to influence various aging processes (74), also has been shown to diminish the age-associated rise in serum IL-6 (75).
IL-6 may be an important mediator of several infectious and autoimmune diseases. These include human immunodeficiency virus (76, 77), rheumatoid arthritis (78), Castleman's disease (79, 80), and the paraneoplastic symptoms associated with cardiac myxoma (81-83). Furthermore, elevated serum and cerebrospinal fluid levels of IL-6 can be found in sepsis (84, 85). Inflammatory joint disease, particularly rheumatoid arthritis (78), is associated with increased synovial fluid levels of IL-6 (86).
In spite of the great variety of health consequences associated with IL-6, it manifests its activity by binding to a specific receptor, the IL-6R, which is described below.
Human IL-6 has a molecular weight of between 21 to 28 Kd depending on post-translational processing such as glycosylation and phosphorylation (87, 88). The IL-6 peptide contains 212 amino acids (aa) of which a 28 aa hydrophobic signal peptide is cleaved off resulting in a mature protein of 184 aa. Even though the homology between human and mouse IL-6 is 65% at the nucleic acid level and only 42% at the amino acid level (89), human IL-6 can stimulate murine IL-6 responsive cells. This may be due to the highly conserved central region (57% homology at the amino acid level) of the molecule which contains four cysteine residues that can be perfectly aligned between mouse and human IL-6 (90). Additionally, the carboxy-terminus appears to be critical for IL-6 activity (91, 92). When just four aa were deleted from the carboxy-terminus, IL-6 activity was completely lost (93). In contrast, deletion of 28 aa from the amino-terminus did not affect IL-6 activity (94).
The human IL-6 gene, located on chromosome 7p21 (95-97), is approximately 5 Kb (compared to 7 Kb for the mouse (98)) and consists of four introns and five exons (99). The human IL-6 gene contains three transcriptional initiation sites which correspond with three TATA-like sequences (99).
Characterization of the IL-6 gene 5' flanking region has revealed a very complex control region. The importance of this region is underscored by the observation that the proximal 300 bp of the human and murine IL-6 gene 5'-flanking region share approximately 80% homology (98). Figure 3 summarizes the regulatory elements in the IL-6 promoter. Table 1 and Table 2 summarize factors which induce or repress the IL-6 promoter, respectively.
Figure 3: Schematic representation of the IL-6 promoter. See text for abbreviation definitions.
Briefly, several cis-acting response elements mediate activation of the IL-6 promoter including those for AP-1, nuclear factor IL-6 (NF-IL6), NF-kappaB, and the multiple response element (MRE), The MRE and NF-IL6 response element are components of the serum response element (SRE). The SRE was first identified in c-fos and induces gene transcription when serum-starved cells are exposed to serum (122, 123). The MRE confers induction of the IL-6 promoter to TPA, serum, forskolin, IL-1alpha, and TNF (115). Repression of the IL-6 promoter can be mediated by various combinations of trans-acting factors and cis-acting elements including Fos binding to the SRE, retinoblastoma protein binding to the retinoblastoma control element (RCE), and a variety of steroids (described below)