Critical Cytoplasmic Domains of Human Interleukin-9 Receptor α Chain in Interleukin-9-mediated Cell Proliferation and Signal Transduction

Abstract

Interleukin-9 receptor (IL-9R) complex consists of a ligand-specific ␣ chain and IL-2R ␥ chain. In this study, two regions in the cytoplasmic domain of human IL-9R␣ were found to be important for IL-9-mediated cell growth. A membrane-proximal region that contains the BOX1 consensus sequence is required for IL-9-induced cell proliferation and tyrosine phosphorylation of Janus kinases (JAKs). Deletion of this region or internal deletion of the BOX1 motif abrogated IL-9-induced cell proliferation and signal transduction. However, substitution of the Pro-X-Pro in the BOX1 motif with Ala-X-Ala failed to abolish IL-9-induced cell proliferation but decreased IL-9-mediated tyrosine phosphorylation of JAK kinases, insulin receptor substrate-2, and signal transducer and activator of transcription 3 (STAT3) and expression of c-myc and junB. Another important region is downstream of the BOX1 motif and contains a STAT3 binding motif YLPQ. Deletion of this region significantly impaired IL-9-induced cell growth, activation of JAK kinases, insulin receptor substrate-2, and STAT3 and expression of early response genes. A point mutation changing YLPQ into YLPA greatly reduced IL-9-induced activation of STAT3 and expression of c-myc but did not affect cell proliferation. These results suggest that cooperation or cross-talk of signaling molecules associated with different domains of IL-9R␣ other than STAT3 is essential for IL-9-mediated cell growth. Interleukin 9 (IL-9)1 is a T cell-derived multifunctional growth factor that exerts its effects on activated T cells, B cells, mast cells, and hematopoietic progenitors (1, 2). The in vitro biological functions of IL-9 include its ability to stimulate proliferation of activated T cells, to enhance production of immunoglobulin in B cells, and to promote proliferation and differentiation of mast cells and hematopoietic progenitors (3-7). The involvement of IL-9 in lymphomagenesis has been suggested by in vivo studies (8) in which a higher susceptibility to T cell lymphoma was observed in transgenic mice expressing IL-9 constitutively and by in vitro experiments (9) in which IL-9 has been shown to protect mouse lymphoma cells from dexamethasone-induced apoptosis. The functions of IL-9 are mediated by the IL-9 receptor (IL-9R), which consists of a ligand-specific ␣ chain and IL-2 receptor (IL-2R) ␥ chain. IL-2R ␥ chain, normally referred to as the common ␥ chain (␥ c chain), is shared by receptors for IL-2, IL-4, IL-7, IL-9, and IL-15 (10 -13). Some of the signaling pathways elicited by these cytokines are quite similar, which probably explains in part the functional redundancy of these cytokines.IL-9R ␣ chain (IL-9R␣) is IL-9-specific and is responsible for IL-9 binding. The cDNAs encoding mouse and human IL-9R␣ have been cloned (14 -15). IL-9R␣ belongs to the hematopoietic receptor superfamily and has no intrinsic tyrosine kinase motif in its cytoplasmic region. Several homologous sequences, such as the BOX1 consensus sequence and a serine-rich region, which were demonstrated to be import...

IL-9-Mediated Induction of Eotaxin1/CCL11 in Human Airway Smooth Muscle Cells

“…Previous studies have shown a constitutive association of JAK-1 with the IL-9R, and noted the presence of IL-9-induced activated complexes containing STAT-1, STAT-3, and STAT-5 (49,50). Moreover, other studies have shown that IL-9 induced tyrosine phosphorylation of insulin receptor substrate-2 through JAK kinases activity (51). In tracheal ASM cells, the ERK/MAPK pathway has recently been shown to be involved in IL-9 activation (39).…”

Regulation of Jak Kinases by Intracellular Leptin Receptor Sequences

“…Indeed, receptors that predominantly signal via Jak1 and/or Tyk2 (such as gp130, IFNR␣, and IL2R␤) do not contain this motif and generally require sequences COOHterminal to this region (e.g. Box 2) for association with/signaling by these Jak kinase isoforms (31,33). In contrast, as for LRb, the residues that make up this (E/N/Q)-rich motif in the Epo receptor are required for Jak2 association, whereas residues immediately COOH-terminal to this motif are not required (27).…”

Involvement of the Ubiquitin-Proteasome Pathway in the Degradation of Nontyrosine Kinase-Type Cytokine Receptors of IL-9, IL-2, and Erythropoietin

“…To differentiate between these two possibilities, we used a panel of TS1 cell lines stably transfected with IL-9R␣ mutants (21) (summarized in Fig. 7) to examine whether any cytoplasmic domains of IL-9R␣ are critical for VCP interaction.…”

“…TS1 cells stably transfected with human IL-9R␣ variants (21) were cultured in the same medium supplemented with 1 mg/ml G418 (Sigma, St. Louis, MO). For cytokine stimulation, cells were cultured in medium without IL-9 and serum for at least 15 h, then collected and stimulated with 30 ng/ml recombinant human IL-9.…”