We show that co-expression of interleukin 15 (IL-15) and IL-15 receptor ␣ (IL-15R␣) in the same cell allows for the intracellular interaction of the two proteins early after translation, resulting in increased stability and secretion of both molecules as a complex. In the absence of co-expressed IL-15R␣, a large portion of the produced IL-15 is rapidly degraded immediately after synthesis. Co-injection into mice of IL-15 and IL-15R␣ expression plasmids led to significantly increased levels of the cytokine in serum as well as increased biological activity of IL-15. Examination of natural killer cells and T lymphocytes in mouse organs showed a great expansion of both cell types in the lung, liver, and spleen. The presence of IL-15R␣ also increased the number of CD44 high memory cells with effector phenotype (CD44 high CD62L؊). Thus, mutual stabilization of IL-15 and IL-15R␣ leads to remarkable increases in production, stability, and tissue availability of bioactive IL-15 in vivo. The in vivo data show that the most potent form of IL-15 is as part of a complex with its receptor ␣ either on the surface of the producing cells or as a soluble extracellular complex. These results explain the reason for coordinate expression of IL-15 and IL-15R␣ in the same cell and suggest that the IL-15R␣ is part of the active IL-15 cytokine rather than part of the receptor. 2 is a pleiotropic cytokine produced in many tissues. It is a member of the four ␣-helix bundle family of cytokines and was initially described as a T cell proliferation factor (1, 2). IL-15 shares with interleukin-2 (IL-2) a common receptor complex, consisting of the IL-2 receptors  and ␥ chains (3). Both IL-2 and IL-15 use an additional private receptor subunit responsible for the specificity of binding, the IL-2 receptor ␣ (IL-2R␣) and IL-15 receptor ␣ (IL-15R␣), respectively. Both molecules have a similar ligand-binding motif (sushi domain) as well as a relatively short intracellular tail (13 amino acids for human IL-2R␣ and 41 amino acids for human IL-15R␣). In contrast to IL-2R␣, which displays a lower affinity for IL-2 (K d ϳ 10 Ϫ8 M) and is expressed mainly on activated T cells, IL-15R␣ has a high affinity for IL-15 (K d ϳ10 Ϫ11 M), and its mRNA has a wide tissue distribution (4). IL-15Ϫ/Ϫ and IL-15R␣ Ϫ/Ϫ mice have profound defects in NK, NK-T, intraepithelial lymphocytes, and memory CD8ϩ T cells, indicating that IL-15 is essential for the homeostatic maintenance and function of these cells (5, 6). In contrast, IL-2 Ϫ/Ϫ and IL-2R␣ Ϫ/Ϫ mice develop autoimmune diseases with increased frequency of activated T and B cells (7,8). Despite the clear results on the positive role of IL-15R␣ for IL-15 function, several investigators have reported inhibitory effects of IL-15R␣ on IL-15 function. Injection in mice of a soluble recombinant form of IL-15R␣ protein (IL-15sR␣) was reported to suppress natural killer (NK) cell proliferation and T-dependent immune responses in vivo (9). Addition of IL-15sR␣ in vitro was reported to block the response of cell lines to IL-1...
IntroductionIL-15 is a member of the common ␥-chain family of cytokines and was initially characterized as a T-cell proliferation factor. 1,2 IL-15 is a growth, mobilization, and activation factor for important lymphocyte populations, including NK, CD8, and intraepithelial lymphocytes. 3-6 IL-15 and IL-2 share the same IL-2/IL-15␥ receptor (IL-2/IL-15R␥), 7 but their biologic effects at the level of organism are different, as shown by studies in knockout mice. The lack of IL-15 in vivo results in severe defects in the development and function of the immune system. 8,9 Although lymphocytes from IL-2 knockout mice fail to proliferate in response to polyclonal T-cell mitogens in vitro, 10 the common features developing in these mice are lymphocyte activation and autoimmunity. 11 In humans, IL-2-receptor ␣ (IL-2R␣) deficiency has been linked to a paradoxical combination of immunodeficiency and autoimmunity in 2 patients. 12,13 The biologic differences of IL-2 and IL-15 are determined by their different production sites, 5,14 their strength of association with the membrane proteins IL-2R␣ and IL-15R␣, 15 respectively, and the regulation of these receptor molecules. IL-2 is produced by activated lymphocytes, whereas IL-15 is produced by stromal cells of several tissues, and by antigen-presenting cells. IL-15R␣ has a high affinity for IL-15 (K d ϳ 10 Ϫ11 M). 15 In contrast, IL-2R␣ has lower affinity for IL-2 (K d ϳ 10 Ϫ8 M) and is expressed mainly on activated T cells and persistently on Treg.IL-15 is known to act on the surface of the cell in complex with IL-15R␣ to engage the IL-2/IL-15R␥ complex in nearby cells, a process termed trans-presentation. 16 Genetic and cell transfer experiments have shown that IL-15 and IL-15R␣ need to reside in the same cell for appropriate function. [17][18][19][20] We previously demonstrated that coexpression of the 2 molecules in the same cell leads to rapid intracellular association of IL-15 and IL-15R␣ in the endoplasmic reticulum, stabilization of both molecules, and the generation of a stable complex that can translocate to the cell membrane, where it is bioactive. 21,22 In addition, the surface heterodimer is rapidly cleaved and released in the plasma as bioactive cytokine. 22 Our experiments using IL-15 complexed to a C-terminal deletion of IL-15R␣ containing only the soluble extracellular fragment demonstrated that this complex is bioactive in vivo in the absence of any membrane-bound form of IL-15. 22 These results explain the necessity for the coordinate expression of IL-15 and IL-15R␣ in the same cell for physiologic function, [17][18][19] and predict that the circulating form of IL-15 in biologic fluids is in complex with soluble IL-15R␣ (sIL-15R␣).Although these results suggested that the main bioactive form of IL-15 is in a complex with IL-15R␣, 21-26 the actual form of IL-15 produced in humans has not been identified. The levels of IL-15 in normal human serum are close to the limit of detection of the existing assay (ϳ 1 pg/mL). Therefore, we studied the nature of the...
We have previously shown that macaques vaccinated with DNA vectors expressing SIVmac239 antigens developed potent immune responses able to reduce viremia upon high-dose SIVmac251 challenge. To further improve vaccine-induced immunity and protection, we combined the SIVmac239 DNA vaccine with protein immunization using inactivated SIVmac239 viral particles as protein source. Twenty-six weeks after the last vaccination, the animals were challenged intrarectally at weekly intervals with a titrated dose of the heterologous SIVsmE660. Two of DNA-protein coimmunized macaques did not become infected after 14 challenges, but all controls were infected by 11 challenges. Vaccinated macaques showed modest protection from SIVsmE660 acquisition compared with naïve controls (P = 0.050; stratified for TRIM5α genotype). Vaccinees had significantly lower peak (1.6 log, P = 0.0048) and chronic phase viremia (P = 0.044), with 73% of the vaccinees suppressing viral replication to levels below assay detection during the 40-wk follow-up. Vaccine-induced immune responses associated significantly with virus control: binding antibody titers and the presence of rectal IgG to SIVsmE660 Env correlated with delayed SIVsmE660 acquisition; SIV-specific cytotoxic T cells, prechallenge CD4 + effector memory, and postchallenge CD8 + transitional memory cells correlated with control of viremia. Thus, SIVmac239 DNA and proteinbased vaccine protocols were able to achieve high, persistent, broad, and effective cellular and humoral immune responses able to delay heterologous SIVsmE660 infection and to provide long-term control of viremia. These studies support a role of DNA and protein-based vaccines for development of an efficacious HIV/AIDS vaccine.T he use of a combination vaccine consisting of the recombinant Canarypox ALVAC-HIV (vCP1521; containing Gag, PR, and Env) together with gp120 Env protein (AIDSVAX B/E) resulted in modest, but statistically significant protection from infection in the RV144 vaccine trial conducted in Thailand (1). The limited efficacy and the fact that the vaccine-induced responses waned over time suggest that improved vaccine designs are needed to achieve long-lasting cross-clade-specific immune responses able to prevent infection. Rhesus macaque simian immunodeficiency virus (SIV) challenge models provide an excellent system to test different vaccine modalities and to compare efficacy using different challenge viruses and infection routes.DNA as priming immunization together with boosting by recombinant viral vectors is a vaccine platform widely used in the HIV/SIV field. DNA as the only vaccine component has been considered poorly immunogenic in humans, although recent results showed that in vivo DNA electroporation (EP) results in more efficient vaccine delivery, a higher frequency of responders, and higher, longer-lasting immunity than needle/syringe delivery (2). Similarly, the inclusion of DNA encoding the cytokine IL-12 as molecular adjuvant has been shown to be advantageous (3). These recent data suggest that DN...
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