Distinct Regions of the IL-7 Receptor Regulate Different Bcl-2 Family Members

Jiang Q, Li WQ, Hofmeister RR, Young HA, Hodge DR, Keller JR, Khaled AR, and Durum SK. Distinct regions of the interleukin-7 receptor regulate different Bcl2 family members. Mol Cell Biol 24: 6501–13, 2004.

nterleukin 7 (IL-7) is a cytokine produced by the non-lymphoid cells in lymphoid tissues. T lymphocytes require IL-7 as they develop in the thymus. After leaving the thymus, they require IL-7 in peripheral sites. This IL-7 requirement is very stringent, such that congenital defects in components of the IL-7 receptor complex are the most frequent cause of severe combined immunodeficiency.

Our lab and others have shown that a major function of IL-7 is to keep the T cell alive. We also showed that this “trophic” function is largely based on the balance of Bcl-2 family proteins that control mitochondrial integrity. Thus, IL-7 induced the synthesis of Bcl-2 and Mcl-1, which protect mitochondria. Withdrawal of IL-7 interrupted their synthesis but also posttranslationally activated the death proteins Bax and Bad, which attack mitochondria.

What is the IL-7 receptor signaling pathway leading to these mitochondrial events? The IL-7 receptor complex has been known for some time to include the IL-7 receptor-α (IL-7Rα) chain, which binds IL-7 avidly, and the γ_c chain, which is recruited to this complex. These two receptor chains each bear kinases on their intracellular domains, Jak1 and Jak3, and when these kinases are brought together, they mutually phosphorylate one another and greatly increase their enzymatic activity. However, we do not know their relevant substrates. We know the initial signaling complex. We also know that the signaling pathway ends with survival proteins, but we do not understand the middle part.

Whatever signaling pathways emanate from the IL-7 receptor, they are surely important pathways in lymphocytes, and they may well go awry in lymphoma. Several signaling pathways have previously been shown to be activated by IL-7, but none have been shown to be essential. For example, stat5 is activated by IL-7, but its deletion in mice did not have the phenotype of IL-7 insufficiency, which would be manifested as a block in thymic development. The PI3K-AKT pathway and src family kinases are also activated by IL-7, but again, knockout mice have not validated their critical roles. The intracellular domain of IL-7Rα presumably acts as a docking site for the first proteins in the signaling cascade. We hoped that by first identifying the essential intracellular domains of the IL-7Rα chain, we could then find the proteins that interact with these domains and, from there, construct the signaling pathway. Our article (Jiang Q et al. Mol Cell Biol 24: 6501–13, 2004) describes our effort to find these critical regions of the IL-7Rα chain.

We first studied IL-7 signals in an IL-7–dependent thymocyte that we had generated from a p53–/– mouse and later verified the principles in T-cell development in vivo. Chimeric receptors were produced using the intracellular domain of mouse IL-7Rα coupled to the extracellular domain of human IL-4Rα. This chimeric receptor conferred a vigorous cellular response to human IL-4 that mimicked the response to IL-7, including regulating Bcl-2, Bax, and Bad. The intracellular domain was then mutated to determine the critical signaling regions, which we narrowed down to two very small sites, Box 1 and Y449 (Figure 1—Jak1 is bound to Box 1).

Figure 1. Model showing the interleukin 7 (IL-7) receptor and the survival and proliferation pathways that emanate from it. The major pathway requires Y449 on the intracellular domain of the IL-7 receptor-α (IL-7Rα) chain. Y449 is phosphorylated by Janus kinases Jak1 and Jak3 and then presumably serves as a docking site for unknown adaptors, termed “X” and “Y.” These initiate signaling, together with stat5, to the transcription of bcl-2, the constraint of Bax to the cytosol, and cell proliferation. A minor signaling pathway is independent of Y449 and, through an unknown kinase “Z,” leads to phosphorylation and sequestration of Bad.

As shown in the figure, we propose that Jak1 initiates two distinct pathways. The major pathway begins with phosphorylation of Y449, which probably serves as a docking site for proteins (yet to be identified) that lead to survival and proliferation. Stat5 docks to Y449 and is phosphorylated and released, then translocates to the nucleus and induces genes. Stat5 augments signaling by increasing Bcl-2 synthesis; however, stat5 is neither necessary nor sufficient. A minor signaling pathway does not require Y449, but leads from Jak1 to inactivation of Bad, and also to the activation of stat1 and stat3.

To evaluate Box 1 and Y449 in T-cell development, we introduced mutated receptors into IL-7Rα–/– hematopoietic stem cells, transferred these stem cells into mice, and observed the effect on thymic development. This confirmed that the major signaling pathway requires Box 1 and Y449, and that a minor pathway is independent of Y449.

We are now seeking to identify the hypothetical proteins (X and Y) that bind Y449, and we are looking for the missing kinase (Z) that phosphorylates Bad at S112 (as described in Li et al. J Biol Chem 279: 29160–6, 2004), that is, “the end of the beginning” and “the beginning of the end” (Winston Churchill).

Scott Durum, PhD
Principal Investigator
Laboratory of Molecular Immunoregulation
NCI-Frederick, Bldg. 560/Rm. 31-71
Tel: 301-846-1545
Fax: 301-846-7042
durums@mail.ncifcrf.gov