Frizzled
Frizzled (FZD) proteins constitute a family of seven-transmembrane-spanning receptors that primarily function as the core receptors for Wnt ligands in metazoans, playing essential roles in diverse cellular processes such as proliferation, differentiation, and polarity establishment.[1] In humans and other vertebrates, there are ten Frizzled genes (FZD1–10), encoding proteins typically comprising 500–700 amino acids, featuring an extracellular cysteine-rich domain (CRD) for ligand binding, seven hydrophobic transmembrane helices characteristic of G protein-coupled receptors (GPCRs), and an intracellular C-terminal tail for signal transduction.[1][2] These receptors are phylogenetically grouped into five subgroups (FZD1/2/7, FZD3/6, FZD4, FZD5/8, and FZD9/10), with structural variations including N- and O-linked glycosylation sites and motifs for post-translational modifications like palmitoylation and phosphorylation that regulate their activity and trafficking.[2][3] Frizzled receptors mediate Wnt signaling through both canonical and non-canonical pathways, with the canonical Wnt/β-catenin pathway being the most studied. In this pathway, Wnt ligands bind to the CRD of Frizzled with high affinity (Kd 1–10 nM), inducing the formation of a ternary complex with co-receptors LRP5/6, which recruits the adaptor protein Dishevelled (DVL) and inhibits the β-catenin destruction complex, leading to β-catenin stabilization, nuclear translocation, and activation of target gene transcription.[3] Non-canonical pathways, including planar cell polarity (PCP) and Wnt/Ca²⁺ signaling, involve Frizzled interactions with additional effectors like ROR or RYK co-receptors, regulating cytoskeletal dynamics and calcium release without β-catenin involvement.[1] Frizzled signaling is tightly regulated by ubiquitination (e.g., via RING finger proteins ZNRF3/RNF43), endocytosis, and feedback mechanisms to prevent aberrant activation.[2] Beyond development, where Frizzled receptors are indispensable for embryonic patterning, organogenesis, and neural circuit formation—as evidenced by phenotypes in model organisms like Drosophila (four Fz genes) and C. elegans (four Cfz genes)—dysregulated Frizzled activity contributes to pathologies such as developmental disorders and cancers.[1][4] For instance, mutations in FZD4 cause familial exudative vitreoretinopathy, while overexpression or aberrant Wnt-Frizzled signaling drives tumorigenesis in colorectal, breast, and other cancers by promoting uncontrolled proliferation and metastasis.[1][2] Ongoing research highlights Frizzled as therapeutic targets, with inhibitors like monoclonal antibodies targeting specific subtypes showing promise in reducing tumor growth in preclinical models.[2]Discovery and Overview
Historical Discovery
The frizzled (fz) gene was initially identified in Drosophila melanogaster in 1989 as a genetic locus required for planar cell polarity in epidermal structures, such as the orientation of bristles and hairs on the wing and abdomen.[5] This discovery revealed that the gene encodes a protein with seven potential transmembrane domains, suggesting a role in cell surface signaling.[6] In the early 1990s, researchers cloned the first mammalian homologs of frizzled, marking the expansion of the family beyond invertebrates. Specifically, FZD1 and FZD2 were isolated in 1992 from a rat osteosarcoma cell cDNA library, demonstrating sequence similarity to the Drosophila protein and widespread expression in tissues like kidney, liver, heart, uterus, and ovary.[7] Between 1996 and 1998, frizzled proteins were recognized as receptors for Wnt signaling ligands, with studies establishing their functional links to downstream components like Dishevelled and the stabilization of β-catenin.[8] For instance, in 1996, a Drosophila frizzled homolog was shown to act as a receptor for Wingless, the fly ortholog of vertebrate Wnt proteins, thereby initiating polarity and patterning signals.[9] These findings positioned frizzled as central mediators in Wnt-dependent processes, such as embryonic development and tissue homeostasis. A influential 2004 review synthesized these advances, classifying frizzled receptors as atypical G-protein-coupled receptors capable of transducing diverse signals beyond canonical Wnt pathways.Definition and General Characteristics
Frizzled receptors constitute a family of atypical G protein-coupled receptors (GPCRs) classified within the class F subfamily, comprising 10 members in humans designated FZD1 through FZD10.[10] These receptors are characterized by a canonical seven-transmembrane domain topology typical of GPCRs, yet they diverge from classical family A and B GPCRs by lacking conserved motifs such as the DRY sequence at the end of transmembrane helix 3, which is essential for conventional G protein coupling in many GPCRs.[11] Instead, Frizzled receptors primarily transduce signals through interactions with intracellular scaffold proteins rather than direct heterotrimeric G protein activation, although emerging evidence indicates selective G protein involvement in certain contexts.[11] The primary function of Frizzled receptors is to serve as cell surface receptors for secreted Wnt glycoproteins, a family of lipid-modified signaling molecules that play pivotal roles in embryonic development, tissue homeostasis, and stem cell regulation.[12] Upon Wnt ligand binding, Frizzled receptors recruit and activate the intracellular adaptor protein Dishevelled (DVL), which initiates downstream signaling cascades.[12] This activation is facilitated by the receptors' general topological architecture, which includes an N-terminal extracellular cysteine-rich domain (CRD) responsible for ligand recognition, a bundle of seven-pass transmembrane helices that span the plasma membrane to propagate the signal, and a C-terminal intracellular tail that interacts with cytosolic effectors like DVL. A hallmark of the Frizzled family is the high conservation of structural features within the CRD, particularly the presence of 10 invariant cysteine residues across all members, which form five intramolecular disulfide bonds to stabilize the domain's fold and enable high-affinity binding to Wnt ligands.[10] This CRD, approximately 120 amino acids in length, adopts a compact β-sheet-rich structure that presents a hydrophobic groove for accommodating the palmitoylated N-terminus of Wnt proteins, underscoring the evolutionary adaptation of Frizzled receptors for specific ligand interactions.[10] The intracellular C-terminal tail, varying in length among isoforms, often concludes with motifs that facilitate assembly of multiprotein signaling complexes, further distinguishing Frizzled from traditional GPCRs in their reliance on non-canonical transduction mechanisms.[12]Molecular Structure
Extracellular Cysteine-Rich Domain
The extracellular cysteine-rich domain (CRD) of Frizzled receptors serves as the primary N-terminal region for Wnt ligand recognition, spanning approximately 120 amino acids and positioned extracellularly to initiate signaling upon binding. This domain is essential for the receptor's function, as it directly interacts with Wnt proteins to activate downstream pathways.[1] Structurally, the CRD features 10 highly conserved cysteine residues that form five intramolecular disulfide bridges, stabilizing a compact β-sheet-rich fold characteristic of the Frizzled family. This architecture creates a positively charged electrostatic surface on one face of the domain, which facilitates interaction with the lipid-modified N-terminal palmitate group of Wnt ligands. The disulfide bonds ensure structural integrity, with mutagenesis studies confirming their role in maintaining Wnt-binding competence. Insights from crystallographic analyses, such as the 2001 structure of the mouse Frizzled-8 CRD (PDB: 1IJ7), reveal a compact domain with exposed hydrophobic grooves that accommodate the palmitoylated thumb loop of Wnt proteins. Subsequent complex structures, including the 2012 Xenopus Wnt8–mouse Frizzled-8 CRD assembly (PDB: 4F0A), demonstrate how these grooves enable specific insertion of the Wnt lipid moiety, promoting high-affinity binding with dissociation constants in the nanomolar range. These structural details highlight the CRD's role in ligand specificity, independent of the receptor's transmembrane regions. The CRD also confers pathway selectivity among Frizzled subtypes; for instance, the Frizzled-8 CRD exhibits a preference for canonical ligands like Wnt3a, facilitating β-catenin-dependent signaling over non-canonical routes. This selectivity arises from variations in the CRD's binding pocket, allowing differential engagement of Wnt isoforms despite overall sequence conservation across the family.Transmembrane and Intracellular Regions
The transmembrane domain of Frizzled receptors consists of seven alpha-helices that span the plasma membrane, forming a 7TMR (seven-transmembrane receptor) core characteristic of class F G protein-coupled receptors (GPCRs).[2] This heptahelical bundle is atypical among GPCRs due to the absence of the conserved DRY motif at the cytoplasmic end of transmembrane helix 3, which is essential for classical G protein activation in class A GPCRs.[13] Instead, Frizzleds primarily initiate signaling through recruitment of the cytosolic scaffold protein Dishevelled, bypassing direct G protein coupling in the canonical Wnt pathway.[14] The intracellular regions include three cytoplasmic loops and a C-terminal tail, with the latter varying in length from approximately 30 to 150 residues across Frizzled family members.[1] This tail is largely intrinsically disordered, contributing to its flexibility, and contains a highly conserved KTxxxW motif near its end that binds the PDZ domain of Dishevelled, facilitating downstream signal transduction.[15] Mutations in this motif disrupt Dishevelled recruitment and impair Wnt signaling.[16] Recent cryo-EM structures from the 2020s, such as those of human Frizzled-5, Frizzled-7, and Frizzled-8 in complex with Wnt ligands or effectors, have resolved the transmembrane helices at resolutions around 3.5–4 Å, revealing a compact bundle with specific lipid interactions stabilizing the core, including cholesterol and phospholipid binding sites.[17][18] More recent examples include the 2024 cryo-EM structure of FZD4 with the DEP domain of Dishevelled 2 at 3.2 Å resolution, elucidating effector binding interfaces, and a high-resolution (1.9 Å) structure of inactive FZD7, providing details on allosteric regulation.[14][19] However, the full intracellular domains, particularly the flexible C-terminal tail, remain unresolved in these models due to their dynamic nature.[20] Upon Wnt binding to the extracellular cysteine-rich domain, the transmembrane helices undergo conformational rearrangements, such as outward tilting of helix 6, to expose intracellular interfaces for effector recruitment and signal initiation.[20]Classification and Evolution
Frizzled Family Members in Humans
In humans, the Frizzled (FZD) family consists of ten isoforms, FZD1 through FZD10, encoded by distinct genes and classified as class F G-protein-coupled receptors that primarily mediate Wnt signaling.[21] These receptors share overall sequence identities of 20-40% across the family, with greater conservation (50-80%) within phylogenetic subgroups: FZD1/FZD2/FZD7 (approximately 75% identity), FZD3/FZD6 (approximately 53% identity), FZD4/FZD9/FZD10 (approximately 65% identity), and FZD5/FZD8 (approximately 70% identity); the cysteine-rich domain (CRD) and transmembrane regions exhibit the highest conservation among all isoforms.[22][23] Ligand binding shows isoform specificity, with Wnt proteins interacting promiscuously but preferentially; for example, FZD1, FZD2, and FZD7 commonly bind non-canonical ligand Wnt5a, while FZD5 and FZD8 preferentially engage canonical ligand Wnt3a.[13] The chromosomal locations of the FZD genes are dispersed across multiple chromosomes, reflecting their evolutionary divergence. Tissue expression patterns vary, with many isoforms prominently active in neural, cardiovascular, and reproductive tissues during development and adulthood, though some show restricted or low basal expression in normal tissues. Below is a summary of key features for each human FZD isoform:| Isoform | Chromosomal Location | Representative Normal Tissue Expression | Isoform-Specific Ligands (Examples) |
|---|---|---|---|
| FZD1 | 7q21.13 | Brain (cerebral cortex), heart (fetal), placenta | Wnt1, Wnt3a, Wnt5a [13] |
| FZD2 | 17q21.31 | Widely expressed in adult and fetal tissues, including heart, lung, and kidney | Wnt2, Wnt3a, Wnt5a [13] |
| FZD3 | 8p21 | Neural tissues (brain, spinal cord) | Wnt2, Wnt3a, Wnt5a [13] |
| FZD4 | 11q14.2 | Retina, vascular endothelium | Wnt2, Wnt5a, Norrin (non-Wnt ligand) [13] |
| FZD5 | 2q33.3 | Placenta, heart, lung | Wnt2, Wnt3a, Wnt5a [13] |
| FZD6 | 8q22.3 | Skin, brain, kidney | Wnt3a, Wnt5a [13] |
| FZD7 | 2q33.1 | Placenta, intestinal epithelium | Wnt3a, Wnt5a, Wnt7a [13] |
| FZD8 | 10p11.21 | Kidney, pancreas, brain | Wnt1, Wnt3a [13] |
| FZD9 | 7q11.23 | Brain, testis, skeletal muscle | Wnt2 [13] |
| FZD10 | 12q24.33 | Placenta, brain, heart, lung, skeletal muscle | Wnt3a, Wnt7b [13] |