Chemokine CXCL12 Induces Binding of Ferritin Heavy Chain to the Chemokine Receptor CXCR4, Alters CXCR4 Signaling, and Induces Phosphorylation and Nuclear Translocation of Ferritin Heavy Chain

Abstract

Chemokine receptor-initiated signaling plays critical roles in cell differentiation, proliferation, and migration. However, the regulation of chemokine receptor signaling under physiological and pathological conditions is not fully understood. In the present study, we demonstrate that the CXC chemokine receptor 4 (CXCR4) formed a complex with ferritin heavy chain (FHC) in a ligand-dependent manner. Our in vitro binding assays revealed that purified FHC associated with both the glutathione S-transferase-conjugated N-terminal and C-terminal domains of CXCR4, thereby suggesting the presence of more than one FHC binding site in the protein sequence of CXCR4. Using confocal microscopy, we observed that stimulation with CXCL12, the receptor ligand, induced colocalization of the internalized CXCR4 with FHC into internal vesicles. Furthermore, after CXCL12 treatment, FHC underwent time-dependent nuclear translocation and phosphorylation at serine residues. By contrast, a mutant form of FHC in which serine 178 was replaced by alanine (S178A) failed to undergo phosphorylation, suggesting that serine 178 is the major phosphorylation site. Compared with the wild type FHC, the FHC-S178A mutant exhibited reduced association with CXCR4 and constitutive nuclear translocation. We also found that CXCR4-mediated extracellular signal-regulated kinase 1/2 (ERK1/2) activation and chemotaxis were inhibited by overexpression of wild type FHC but not FHC-S178A mutant, and were prolonged by FHC knockdown. In addition to CXCR4, other chemokine receptor-initiated signaling appeared to be similarly regulated by FHC, because CXCR2-mediated ERK1/2 activation was also inhibited by FHC overexpression and prolonged by FHC knockdown. Altogether, our data provide strong evidence for an important role of FHC in chemokine receptor signaling and receptor-mediated cell migration.Chemokines comprise a large family of low molecular mass (8 -10 kDa) proteins with chemotactic and proactivatory effects on different leukocyte lineages. Approximately 40 chemokines have been identified and several studies have established their central role in a number of physiological situations, including T helper cell responses, hematopoiesis, homeostasis, and angiogenesis (1). Chemokines mediate their biological effects by binding to specific seven-transmembrane domain G protein-coupled receptor subtypes, designated CXCR1 through CXCR6, CCR1 through CCR11, XCR1, and CX3CR1, based on their specific preference for certain chemokines (2, 3). Among these chemokine receptors, CXCR4 has received a great deal of attention because this receptor subtype plays a role not only in leukocyte homing but also in human immunodeficiency virus infection, development of the immune and central nervous systems, and cancer metastasis (4 -9). Its ligand, CXCL12, which also binds to RDC1 that is proposed to rename as CXCR7 (10), has been found to induce migration of peripheral blood lymphocytes (11), CD34ϩ progenitor cells (4), and pre-and pro-B cell lines (12). Mice embryos lacking either the C...

CXCR4 nuclear localization follows binding of its ligand SDF-1 and occurs in metastatic but not primary renal cell carcinoma

“…A recent study found several proteins, including ferritin (19) and Hsc73 (20), in combination with CXCR4 after internalization of CXCR4. The ferritin heavy chain and Hsc73 are not only found as CXCR4 binding proteins but also regulate the receptor endocytosis and the receptor-mediated chemotaxis (19,20). It has also been reported that ß-arrestin may regulate the internalization of CXCR4 (22).…”

“…Internalization of CXCR4 after SDF-1 treatment involves interaction with the proteins ferritin (19), 73-kDa heat shock cognate protein (Hsc73) (20), plectin (10) and Myosin IIA (21). A recent study found several proteins, including ferritin (19) and Hsc73 (20), in combination with CXCR4 after internalization of CXCR4.…”

Acute Phase Proteins: Ferritin and Ferritin Isoforms

“…The third receptor involves the CXCL12 chemokines and their receprtors. It has been shown that H-subunit rich ferritins can repress chemokine receptor-mediated signal transduction and migration of lymphocytes by binding to the CXCL12 chemokine's receptor (Li et al, 2006). The CXCL12 chemokines have chemotactic and various activating effects that play important roles in T-helper cell responses.…”

“…On the inside of the cell the H-subunit rich ferritins are phosphorylated followed by nuclear translocation. Furthermore, intracellular H-subunit rich ferritins can repress CXCR4 activation followed by inhibition of lymphocyte migration (Li et al, 2006). Most of the immunomodulatory studies involving H-subunit rich ferritins seem to be of an anti-inflammatory nature.…”

The cSNP scanning and expression analysis of the duck FTH1 gene

“…Thus, the major function of FTH1 is protecting the cell from oxidative damage (11). Recent data indicated that the FTH1 gene is also associated with several disease processes, such as inflammation, tumor progression, and so on (12,13).…”