CXCR4 The Cellular Pathfinder in Immune Response and Beyond

What is CXCR4？

CXCR4, or C-X-C chemokine receptor 4, is a vital protein receptor that belongs to the family of G protein-coupled receptors and features seven transmembrane domains. This receptor is prominently found on the surface of human cell membranes, playing a pivotal role in various biological processes such as cell migration, localization, and signal transduction.One of the most significant aspects of CXCR4 is its interaction with CXCL12, another protein. This interaction holds immense importance as it regulates critical cellular activities including migration and localization. Within the immune system, CXCL12 assumes a crucial role in several functions. It facilitates the myeloid localization of hematopoietic stem cells, aids in the migration of immune cells, and contributes to the orchestration of inflammatory responses.

The CXCR4-CXCL12 axis stands as a prime example of the intricacies present within molecular interactions. This duo's interplay demonstrates the elegance of how cells communicate and coordinate, resulting in the proper functioning of various physiological mechanisms.

The Structure of CXCR4

CXCR4, a protein integral to cell function, exhibits a complex structure comprising 352 amino acids. This rhodopsin-like G-protein-coupled receptor (GPCR) consists of distinct components, including an extracellular N-terminal domain, seven transmembrane (TM) helices, three extracellular loops (ECL), three intracellular loops (ICL), and an intracellular C-terminal domain[1].

Intriguingly, CXCR4 can manifest in various forms within the plasma membrane—ranging from monomers and dimers to higher-order oligomers and nanoclusters[2]. However, the exact significance of these different multimerization states remains unexplored in vivo. Crystal structures of CXCR4, when bound to agonists and small molecules, align with its capacity to form homodimers via interactions between the TM5 and TM6 helices. TM6 is also implicated in nanoclustering. Additionally, CXCR4 can engage in heterodimerization with ACKR3, a closely related GPCR also known as CXCR7, which offers distinct signaling properties[3].

CXCR4's primary ligand is CXCL12, also recognized as stromal cell-derived factor 1 (SDF-1)[4]. Notably, the human CXCL12 gene codes for six protein isoforms (three in mice), resulting from alternative splicing of the fourth and final exon. These isoforms exhibit differential expression patterns and affinities for glycosaminoglycans present on cell surfaces and within the extracellular matrix[5]. Among these isoforms, CXCL12α, an 89 amino acid protein, stands as the shortest and most abundantly expressed variant[6]. Remarkably, CXCL12α can exist both as a monomer and a dimer. CXCL12 selectively binds solely to two chemokine receptors—CXCR4 and ACKR3, the latter being a CXCR4 interactor. Such a restricted receptor selectivity is an uncommon trait among chemokines.

It is worth noting that while the structure of the CXCR4/CXCL12 complex remains undetermined, a model incorporating homology modeling, experimentally derived restraints, and charge swap mutagenesis[7] highlights several interactions between the N-terminal tail of CXCR4 and CXCL12. Additionally, it emphasizes the involvement of the N-terminus of CXCL12 within the cavity formed by the TM helices.

CXCR4 Expression and Function

Originally recognized as the leukocyte-derived seven-transmembrane domain receptor (LESTR) or Fusin, CXCR4 emerged as an orphan GPCR with a striking role in facilitating HIV-1 fusion with target cells, which led to its initial moniker "Fusin"[9]. Unique in its specificity, CXCL12 serves as the dedicated chemokine for CXCR4[10]. This interaction triggers the activation of heterotrimeric Gαβγ proteins, subsequently igniting an array of signaling pathways that control diverse cellular processes including calcium mobilization, actin polymerization, cytoskeletal rearrangements, gene transcription, and receptor internalization[11]. Astonishingly, these pathways influence not just cell proliferation and survival, but also apoptosis[12].

Remarkably, CXCR4's influence extends widely across both embryonic and adult tissues, emphasizing its role as a homeostatic receptor. Research involving Cxcr4-deficient mice mirrors findings from Cxcl12-knockout mice, both showing disruptions in hematopoiesis, nervous system development, and cardiovascular formation. The significance of the CXCR4/CXCL12 axis is underscored by the fact that both knockout mice exhibit embryonic lethality, highlighting the profound importance of these chemokine and chemokine receptor families throughout evolution[13].

While CXCR4's presence is pervasive within the hematopoietic system, its expression levels can significantly vary among different cell types. In line with other chemokine receptors, CXCR4 plays a critical role in leukocyte trafficking and their anchoring within specific niches, both during normal conditions and under pathological circumstances. During the interaction between antigen-presenting cells and T cells, CXCR4, among other chemokine receptors, gathers at the peripheral SupraMolecular Activation Cluster (pSMAC), aiding in integrin activation essential for productive immunological synapse formation and effective T cell activation[14]. In essence, CXCL12/CXCR4 stand as pivotal players in adaptive and innate immune responses, as well as in bone marrow organization and maintenance. They orchestrate hematopoietic stem cell migration, homing, and survival within the bone marrow[15].

Beyond hematopoiesis, CXCR4 also graces various non-hematopoietic tissues such as the lung, liver, kidney, gastrointestinal tract, adrenal gland, ovary, and brain. Conditional Cxcr4-knockout mice have lent insight into CXCR4's crucial roles in adult tissues, revealing its contributions to central nervous system development, and the formation of vasculature in the gastrointestinal tract and kidney[16].

Although primarily deemed a homeostatic receptor, CXCR4's expression can be modulated by diverse pathological conditions. For instance, many tumor types, including breast, ovarian, prostate[17], melanoma, and neuroblastoma, exhibit heightened CXCR4 expression. Moreover, increased CXCR4 expression in metastatic lesions correlates with advanced tumor progression and the selection of preferential metastatic sites. Mouse studies corroborate CXCR4's significance as a cancer target, as blocking it impedes cancer cell spread and metastasis across various cancer models[18]. The CXCL12/CXCR4 duo also actively participates in tumor growth, the interaction of tumor cells with their microenvironment, and processes like vasculogenesis and angiogenesis. In this complex framework, hypoxia has been linked to heightened CXCR4 expression, suggesting the receptor's involvement in tumor progression[19].

The realm of inflammation also plays a pivotal role in modulating CXCR4. Notably, TGF-β1, VGEF, and bFGF are documented to increase CXCR4 expression, while other cytokines like IL-5[20], IFNα, and IFNγ suppress its expression. Collectively, these insights underscore the profound involvement of the CXCR4/CXCL12 axis in immunodeficiency, inflammatory diseases, and cancer, highlighting its potential as a therapeutic target.

CXCR4 Signaling

When ligands bind to CXCR4, a series of intricate events are set in motion, culminating in the activation of multiple signaling pathways. These pathways are initiated by the dissociation of heterotrimeric G proteins and the phosphorylation of CXCR4's C-terminal cytoplasmic tail.

CXCR4 primarily binds to heterotrimeric Gi proteins, although other classes of G proteins can also mediate CXCR4 signaling. Upon ligand binding, the Gi heterotrimer separates into GTP-bound αi and βγ subunits[21]. The βγ subunits directly activate phosphatidylinositol-3-OH kinases (PI3K) β or γ, generating phosphatidylinositol triphosphate (PIP3), and phospholipase C β (PLC-β), producing inositol-(1,4,5)-trisphosphate (IP3) and diacylglycerol (DAG). Simultaneously, the Gαi subunit triggers calcium release from intracellular stores and indirectly activates the PI3K-AKT and MEK1/2-Erk1/2 axes. Through PIP3 production, PI3Ks activate the serine-threonine kinase AKT, which in turn phosphorylates various target proteins, including glycogen synthase kinase 3 (GSK3), tuberous sclerosis 2 (TSC2), caspase 9, and PRAS40 (AKT1S1). This cascade underpins a wide array of downstream effects, including the promotion of cell proliferation, differentiation, apoptosis, angiogenesis, and metabolism.

Interestingly, CXCR4 ligand binding activates JAK/STAT signaling independently of Gα proteins. GPCR kinases (GRKs) phosphorylate multiple serine and threonine sites in CXCR4's cytoplasmic tail. Phosphorylated CXCR4 recruits β-arrestin-1 and -2, which facilitate CXCR4 internalization[22]. Following internalization, CXCR4 can be either recycled to the plasma membrane or routed to lysosomes for degradation. It's worth noting that the recruitment of β-arrestins to CXCR4 also initiates Erk signaling.Binding CXCL12 to CXCR4-ACKR3 heterodimers triggers G protein-independent signaling cascades initiated by β-arrestins, further enhancing cell migration.The activation of PI3Ks and Akt supports the proliferation and survival of both normal and cancer cells. The activation of mTORC drives anabolic metabolism necessary for cell growth and the transition of stem cells to the GAlert state.Crucially, the pathways activated by CXCR4 that govern cell movement and migration mirror those involved in cell proliferation. Both processes can be inhibited by the same small molecules. For example, rapamycin, an mTORC inhibitor, not only blocks cell proliferation but also inhibits cell migration[23], a characteristic shared by PI3K inhibitors.

Despite our understanding of CXCR4-initiated pathways, the potential for cell-specific variations remains vast. The human genome encodes 18 different Gα proteins, 5 Gβ proteins, and 12 Gγ proteins, along with multiple PI3Ks and PLCs, each with varying expression levels in different cell types. Additionally, signaling is modulated by dozens of regulators, including scaffold proteins that facilitate physical interactions among kinases and other enzymes responsible for post-translational modifications. Currently, studies haven't delved into the granular details of CXCR4-controlled proliferation, down to the specific isoforms and post-translational modifications of the involved signal transducers. The complexity of cancer cells, with their genetic and epigenetic alterations, adds another layer of intricacy. Investigating CXCR4-controlled proliferation following injury using cell-specific conditional mutants may provide a list of components in specific cells. However, mechanistic details such as interactions with modifiers, possible feed-forward and feedback loops, and time-dependent signal adaptations remain uncharted territory, akin to the dynamics observed in pathways involving Rac, NF-κB, and p53[24].

CXCR4 Protein

Recombinant Human CXCR4 Protein

Click here for more CXCR4

Synonym : C-X-C chemokine receptor type 4 CD184 CD184 antigen Chemokine (C X C motif) receptor 4 Chemokine CXC Motif Receptor 4 CXC-R4 CXCR-4 CXCR4 CXCR4_HUMAN D2S201E FB22 Fusin HM89 HSY3RR LAP 3 LAP3 LCR1 LESTR Leukocyte derived seven transmembrane domain receptor Leukocyte-derived seven transmembrane domain receptor Lipopolysaccharide associated protein 3 Neuropeptide Y receptor Y3 NPY3R NPYR NPYRL NPYY3 NPYY3R Probable G protein coupled receptor lcr1 homolog SDF 1 receptor SDF-1 receptor SEVEN-TRANSMEMBRANE-SEGMENT RECEPTOR Stromal cell derived factor 1 receptor Stromal cell-derived factor 1 receptor WHIM WHIMS

References:

 

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