Loss of nuclear pro-IL-16 facilitates cell cycle progression in human cutaneous T cell lymphoma.

Cutaneous T cell lymphomas (CTCLs) represent a heterogeneous group of non-Hodgkin lymphomas that affect the skin. The pathogenesis of these conditions is poorly understood. For example, the signaling mechanisms contributing to the dysregulated growth of the neoplastic T cells are not well defined. Here, we demonstrate that loss of nuclear localization of pro-IL-16 facilitates CTCL cell proliferation by causing a decrease in expression of the cyclin dependent-kinase inhibitor p27Kip1. The decrease in p27Kip1 expression was directly attributable to an increase in expression of S-phase kinase-associated protein 2 (Skp2). Regulation of Skp2 is in part attributed to the nuclear presence of the scaffold protein pro-IL-16. T cells isolated from 11 patients with advanced CTCL, but not those from healthy controls or patients with T cell acute lymphocytic leukemia (T-ALL), demonstrated reduction in nuclear pro-IL-16 levels. Sequence analysis identified the presence of mutations in the 5' end of the PDZ1 region of pro-IL-16, a domain required for association of pro-IL-16 with the nuclear chaperone HSC70 (also known as HSPA8). HSC70 knockdown led to loss of nuclear translocation by pro-IL-16 and subsequent increases in Skp2 levels and decreases in p27Kip1 levels, which ultimately enhanced T cell proliferation. Thus, our data indicate that advanced CTCL cell growth is facilitated, at least in part, by mutations in the scaffold protein pro-IL-16, which directly regulates Skp2 synthesis.

Preferential migration of T regulatory cells induced by IL-16.

As a natural ligand for CD4, IL-16 has been shown to preferentially induce migration in Th1 cells, and, in long-term cultures with IL-2, IL-16 facilitates the expansion of CD4(+)CD25(+) cells. In addition, IL-16 has an immunomodulatory role in asthmatic inflammation, as exogenous administration significantly reduces inflammation and airway hyperreactivity. The mechanism for this, however, is not clear. Based on its functional characteristics and potential immunomodulatory role, we investigated the ability of IL-16 to recruit and influence the development of T regulatory (Treg) cells. We now demonstrate that IL-16 preferentially induces migration in a CD25(+)CTLA-4(+) human T cell subset and that responding cells produce IFNgamma and TGFbeta but not IL-10. These cells are relatively unresponsive to antigenic stimulation and can suppress proliferation and IL-5, but not IFNgamma, production by autologous T cells. We further demonstrate that IL-16-recruited cells are enriched for Forkhead box P3 (Foxp3). In addition, we find that IL-16 stimulation may facilitate de novo induction of Foxp3(+) Treg cells, because the stimulation of FoxP3-negative T cells for 48 h results in the expression of FoxP3 mRNA and protein. These data indicate that at sites of inflammation IL-16 may contribute to selective Treg cell expansion through the preferential induction of a migratory response from existing Treg cells, as well as by the induction of de novo generation of FoxP3(+) cells. These findings offer a potential mechanism for the immunosuppressive effects of IL-16 seen in Th2-mediated inflammation.

Chemokine receptor CXCR3 desensitization by IL-16/CD4 signaling is dependent on CCR5 and intact membrane cholesterol.

Previous work has shown that IL-16/CD4 induces desensitization of both CCR5- and CXCR4-induced migration, with no apparent effect on CCR2b or CCR3. To investigate the functional relationship between CD4 and other chemokine receptors, we determined the effects of IL-16 interaction with CD4 on CXCR3-induced migration. In this study we demonstrate that IL-16/CD4 induced receptor desensitization of CXCR3 on primary human T cells. IL-16/CD4 stimulation does not result in surface modulation of CXCR3 or changes in CXCL10 binding affinity. This effect does require p56(lck) enzymatic activity and the presence of CCR5, because desensitization is not transmitted in the absence of CCR5. Treatment of human T cells with methyl-beta-cyclodextrin, a cholesterol chelator, prevented the desensitization of CXCR3 via IL-16/CD4, which was restored after reloading of cholesterol, indicating a requirement for intact cholesterol. These studies demonstrate an intimate functional relationship among CD4, CCR5, and CXCR3, in which CCR5 can act as an adaptor molecule for CD4 signaling. This process of regulating Th1 cell chemoattraction may represent a mechanism for orchestrating cell recruitment in Th1-mediated diseases.