Modulation of the Interleukin-6 Receptor Subunit Glycoprotein 130 Complex and Its Signaling by LMO4 Interaction

Abstract

The interleukin (IL)-6-type cytokines play major roles in a variety of biological processes by signaling through a common receptor subunit, glycoprotein (gp) 130. We performed yeast two-hybrid screening to identify new binding partners of the activated gp130 and the associated Janus kinases. LMO4, a LIM domain-containing protein that belongs to a family of oncogenes, was identified in this assay. Further studies show that LMO4 associates with gp130 and Janus kinase1 in several mammalian cell types. It also interacts with proteintyrosine phosphatase 2 (SHP2) and suppressor of cytokine signaling 3 (SOCS3). The binding domains involved in these interactions were mapped, and the interactions were shown to be in a direct manner by in vitro binding assays. It is likely that LMO4 exists in the gp130 complex. The cellular localization of LMO4 was detected primarily in the nucleus with a substantial amount also detected in the cytoplasm in several cell types. The interleukin (IL 1 )-6-type cytokines belong to a cytokine family that plays major roles in hematopoiesis, immune responses, inflammation, and neural development by regulating the expression of their target genes (1, 2). IL-6 regulates the production of acute phase proteins in hepatocytes and stimulates differentiation of B and T cells and proliferation of keratinocytes, mesangial, and myeloma/plasmacytoma cells. The IL-6 cytokine family transduces their signals via a common receptor subunit, glycoprotein 130 (gp130). In the case of IL-6, binding of the ligand to the non-signaling ␣-receptor subunit recruits the ␤-receptor subunits, gp130, which form homodimers. The receptor dimerization leads to phosphorylation and activation of the non-receptor tyrosine kinases of the JAK family that are pre-associated with gp130. JAKs in turn phosphorylate specific tyrosine residues in the cytoplasmic portion of gp130, which serve as docking sites for signaling molecules containing Src homology 2 (SH2) domains. Six tyrosine residues are phosphorylated in gp130 upon ligand stimulation, leading to the activation of two major pathways (for viewed, see Ref.3). The four phosphotyrosine residues at the C-terminal region of gp130 serve as recruitment sites for Stat3 (4), whereas phosphorylation of its second membrane-proximal tyrosine residue leads to recruitment of SHP2 (3). SHP2 serves as an adaptor protein mediating the activation of the Ras-Rafmitogen-activated protein kinase signaling pathway. As a protein-tyrosine phosphatase, SHP2 also plays a negative regulatory role in IL-6 signaling (5, 6). The same site also mediates binding of the IL-6-induced feedback inhibitor SOCS3, which attenuates IL-6 signaling by inhibiting JAKs as well as by triggering protein degradation (7,8).The LIM domain, characterized by a double zinc finger structure, was originally identified in the homeodomain transcription factors Lin-11, Isl-1, and Mec-3, which have established roles in the central nervous system, and was subsequently found in a variety of proteins with diverse functions. The LIMcontain...

Cisplatin-induced Ototoxicity Is Mediated by Nitroxidative Modification of Cochlear Proteins Characterized by Nitration of Lmo4

“…Cisplatininduced modulation of Lmo4 can facilitate the inhibition of Stat3 activity by a direct physical interaction of estrogen receptor ␣ with Stat3 (46). Moreover, silencing of Lmo4 expression decreases Stat3 activity (47) as Lmo4 mediates ATP signaling that leads to phosphorylation of Stat3 (48). Because inhibition of Stat3 activity has been implicated in drug-mediated apoptosis (49,50), decreases in cochlear ATP (14) and Lmo4 levels after cisplatin treatment point to the significance of Lmo4 in mediating an apoptotic response.…”

LIM domain only 4 protein promotes granulocyte colony-stimulating factor-induced signaling in neurons

“…LMO4 is primarily localized in the nucleus with low levels also detected in the cytoplasm in several cell types, including hepatocytes and F11 neuronal cells [19,20]. Although LMO4 does not physically interact with Stat3 ( [20], and Chen et al, data not shown), LMO4 interacts with several proteins involved in Stat3 signaling, including the interleukin-6 (IL-6) receptor glycoprotein 130 (gp130), Janus kinase 1 (Jak1), SH2 domain containing tyrosine phosphatase 2 (Shp2) and suppressor of cytokine signaling 3 (SOCS3) [20]. Downregulation of LMO4 shortens gp130 half-life, reduces Stat3 tyrosine phosphorylation and Stat3-dependent gene activation [20].…”

“…Although LMO4 does not physically interact with Stat3 ( [20], and Chen et al, data not shown), LMO4 interacts with several proteins involved in Stat3 signaling, including the interleukin-6 (IL-6) receptor glycoprotein 130 (gp130), Janus kinase 1 (Jak1), SH2 domain containing tyrosine phosphatase 2 (Shp2) and suppressor of cytokine signaling 3 (SOCS3) [20]. Downregulation of LMO4 shortens gp130 half-life, reduces Stat3 tyrosine phosphorylation and Stat3-dependent gene activation [20]. We observed that GCSF treatment increased Stat3 phosphorylation in wild-type but not in LMO4 null cortical neurons, supporting the notion that LMO4 is required for GCSF-induced Stat3 phosphorylation and activation.…”

“…LMO4 was previously reported as a novel regulator of Stat3 signaling in hepatocytes [20]. Over-expression of LMO4 enhanced the transcriptional activity and target gene expression of Stat 3.…”

“…Over-expression of LMO4 enhanced the transcriptional activity and target gene expression of Stat 3. Furthermore, silencing LMO4 expression in stable cell lines expressing small interfering RNA of LMO4 decreased Stat3 activity [20].…”