IL17F Human

What is human IL-17F and what is its relationship to other IL-17 family members?

Human IL-17F is a pro-inflammatory cytokine belonging to the IL-17 family, which includes six members (IL-17A through IL-17F). IL-17F shares the highest amino acid sequence identity (approximately 50%) with IL-17A, making these two the most closely related members of the family . The gene encoding IL-17F is located on chromosome 6, in the same chromosomal region as IL-17A, suggesting they likely arose from gene duplication . IL-17F is synthesized as a 153 amino acid precursor protein with a 20 amino acid signal sequence and a 133 amino acid mature region, and like IL-17A, it contains one potential site for N-linked glycosylation .

Unlike other cytokine families, IL-17 family members have a unique structural fold. Human IL-17F functions as a disulfide-linked homodimer with a molecular weight of approximately 30.1 kDa . Additionally, IL-17F can form heterodimers with IL-17A (IL-17A/F), providing another layer of complexity and functional diversity to this cytokine system .

Which cell types produce IL-17F and under what conditions?

IL-17F is primarily produced by activated CD4+ T cells, particularly those of the Th17 lineage that are stimulated under specific polarizing conditions . When naïve CD4+ T cells are activated in the presence of IL-6 or IL-21 plus TGF-β, they produce high levels of IL-17F, with approximately two-thirds of IL-17F-positive cells co-expressing IL-17A . Interestingly, TGF-β treatment alone can transiently upregulate the expression of IL-17F but not IL-17A at both mRNA and protein levels .

Beyond CD4+ T cells, IL-17F is also produced by:

• γδ T cells

• Invariant natural killer T (iNKT) cells

• Lymphoid-tissue inducer (LTi)-like cells

• Natural killer (NK) cells

• Paneth cells

• Neutrophils

• Potentially activated monocytes, basophils, and mast cells (though protein-level evidence is less clear)

These cellular sources are particularly abundant in barrier tissues such as the skin, gut, and lung, where IL-17F plays important roles in host defense against pathogens .

How does IL-17F signal, and what are its primary biological effects?

IL-17F signals by binding to a receptor complex composed of IL-17RA and IL-17RC . Upon binding, it triggers a signaling cascade that involves Act1 and TRAF6 as signal transducers . This activation leads to the induction of pro-inflammatory cytokines and chemokines in various cell types, including fibroblasts, epithelial cells, and endothelial cells .

The primary biological effects of IL-17F include:

• Induction of pro-inflammatory cytokines (such as IL-6) and chemokines

• Stimulation of proliferation and activation of T-cells and peripheral blood mononuclear cells (PBMCs)

• Regulation of cartilage matrix turnover

• Inhibition of angiogenesis

• Recruitment, activation, and migration of neutrophils

• Protection of skin and mucosal barriers against infectious agents

Importantly, IL-17F appears to have similar but not identical functions to IL-17A, with IL-17F generally demonstrating less potent pro-inflammatory activity in many assays .

What are the key features of the IL-17F crystal structure, and how does it compare to IL-17A?

The crystal structure of human IL-17F reveals a homodimeric cytokine with a unique structural fold characteristic of the IL-17 family . Each monomer contains cysteine knot motifs that are stabilized by intramolecular disulfide bonds, and the dimer is further stabilized by intermolecular disulfide linkages. The interface between the two monomers creates a saddle-shaped topology with receptor binding sites located at the extremities of the dimer .

When comparing IL-17F to IL-17A:

Recent structural studies have shown that IL-17F can form a symmetrical 2:1 complex with IL-17RC (two IL-17RC molecules to one IL-17F homodimer), which is different from the previously expected model of IL-17F signaling .

What is known about the structure of the IL-17A/F heterodimer and its receptor binding properties?

The IL-17A/F heterodimer exhibits a fascinating "two-faced" structure as revealed by X-ray crystallography . One side of the heterodimer closely resembles IL-17A (the "A-face"), while the other side resembles IL-17F (the "F-face"), creating a cytokine with dual binding properties .

Key characteristics of the IL-17A/F heterodimer include:

Interestingly, crystallographic studies have shown that IL-17RA can bind to the "F-face" of the IL-17A/F heterodimer, despite IL-17RA having a much higher affinity for IL-17A. Through site-directed mutagenesis, researchers demonstrated that IL-17RA can also bind to the "A-face" of IL-17A/F with similar affinity . Furthermore, IL-17RC does not discriminate between the two faces of the heterodimer, enabling the formation of two topologically distinct heterotrimeric complexes with potentially different signaling properties .

How do the receptor binding interfaces of IL-17F differ from other IL-17 family members?

The receptor binding interfaces of IL-17F have several distinctive features compared to other IL-17 family members:

• IL-17F binds to both IL-17RA and IL-17RC, but with different affinities than IL-17A. IL-17F has lower affinity for IL-17RA but higher affinity for IL-17RC compared to IL-17A .

• Recent structural studies have revealed that IL-17F can form a symmetrical 2:1 complex with IL-17RC (two IL-17RC molecules to one IL-17F homodimer), which contradicts the previously accepted model. This finding suggests that IL-17RC competes with IL-17RA for cytokine binding .

• Using biophysical techniques, researchers demonstrated that IL-17A and IL-17A/F also form 2:1 complexes with IL-17RC, suggesting the possibility of IL-17RA-independent IL-17 signaling pathways .

• The formation of asymmetric binary and ternary receptor complexes with IL-17F is in sharp contrast to other cytokine families like TGF-β and BMPs, which form symmetrical signaling complexes in a 2:2:2 stoichiometry .

These structural differences in receptor binding likely contribute to the unique biological activities of IL-17F and explain the differential signaling outcomes when compared to other IL-17 family members.

What are the most effective methods for producing recombinant human IL-17F for research applications?

Production of high-quality recombinant human IL-17F typically employs the following methods:

• Expression Systems: Escherichia coli is the most commonly used expression system for producing recombinant IL-17F . The protein is expressed as a disulfide-linked homodimer of two 134 amino acid chains with a molecular weight of approximately 30.1 kDa.

• Purification Protocol:

  • Initial capture using affinity chromatography (often His-tag based systems)

  • Subsequent purification steps may include ion-exchange chromatography and size-exclusion chromatography

  • Purity verification by SDS-PAGE and HPLC analyses (target >95% purity)

• Quality Control: Functional validation through bioactivity assays, including:

  • Ability to induce IL-6 and IL-8 production in target cells

  • Capacity to stimulate PBMC proliferation

  • Confirmation of proper disulfide bond formation using non-reducing SDS-PAGE

• Storage and Handling: Recombinant IL-17F is typically supplied as a sterile filtered white lyophilized powder that should be stored desiccated at -20°C to maintain stability and bioactivity .

For researchers requiring structural studies, specialized approaches may be needed to ensure proper folding and formation of both homodimeric IL-17F and heterodimeric IL-17A/F complexes.

What are the key considerations for designing experiments to study IL-17F signaling pathways?

When designing experiments to study IL-17F signaling pathways, researchers should consider several important factors:

• Receptor Expression Profiling:

  • Verify the expression levels of both IL-17RA and IL-17RC in your experimental system, as both are required for canonical IL-17F signaling

  • Consider the possibility of IL-17RA-independent signaling through IL-17RC, as suggested by recent structural studies

• Signal Transduction Analysis:

  • Include analysis of Act1 and TRAF6 recruitment and activation, as these are key proximal signaling mediators

  • Monitor downstream activation of NF-κB, MAPK, and C/EBP pathways

  • Consider the potential for distinct signaling outputs from IL-17F homodimers versus IL-17A/F heterodimers

• Cytokine/Chemokine Readouts:

  • Measure production of IL-6, IL-8, G-CSF, and chemokines as functional readouts of IL-17F signaling

  • Include both transcript (qPCR) and protein (ELISA) measurements to capture both early and late responses

  • Compare responses to IL-17F, IL-17A, and IL-17A/F to determine relative potencies

• Controls and Validation:

  • Use neutralizing antibodies against IL-17F, IL-17RA, and IL-17RC to confirm specificity

  • Consider using cells from knockout models or CRISPR-mediated deletion of key signaling components

  • Include appropriate positive controls (e.g., TNF-α or IL-1β) for pro-inflammatory responses

• Temporal Considerations:

  • Design time-course experiments to capture both immediate-early (1-2 hours) and delayed (6-24 hours) responses

  • Account for potential feedback regulation by measuring expression of the receptors and negative regulators over time

How can researchers effectively differentiate between the biological activities of IL-17F homodimers versus IL-17A/F heterodimers?

Differentiating between the biological activities of IL-17F homodimers and IL-17A/F heterodimers presents a significant challenge due to their structural similarities and overlapping receptor usage. Here are methodological approaches to address this question:

• Recombinant Protein Production:

  • Generate pure IL-17F homodimers using bacterial expression systems

  • Produce IL-17A/F heterodimers using co-expression systems with differentially tagged IL-17A and IL-17F subunits, followed by tandem affinity purification

  • Verify the identity and purity of these proteins using mass spectrometry and non-reducing SDS-PAGE

• Receptor Binding Assays:

  • Perform surface plasmon resonance (SPR) or bio-layer interferometry (BLI) to measure binding kinetics to IL-17RA and IL-17RC

  • Use competitive binding assays to determine if the homodimers and heterodimers compete for the same binding sites

  • Employ cell-based binding assays with fluorescently labeled cytokines to visualize receptor engagement

• Signaling Analysis:

  • Compare the activation kinetics and magnitude of downstream signaling mediators (e.g., phosphorylation of MAPKs, NF-κB activation)

  • Use reporter cell lines expressing luciferase under the control of responsive promoters (e.g., NF-κB responsive elements)

  • Perform RNA-sequencing to identify differentially regulated genes between homodimer and heterodimer stimulation

• Functional Assays:

  • Compare dose-response curves for induction of IL-6, IL-8, and other inflammatory mediators

  • Analyze neutrophil recruitment and activation in response to each cytokine form

  • Measure effects on epithelial barrier function and antimicrobial peptide production

• Selective Neutralization:

  • Use antibodies that selectively neutralize IL-17A, IL-17F, or both to dissect their relative contributions in complex biological systems

  • Employ receptor mutants that preferentially bind one cytokine form over the other

By combining these approaches, researchers can begin to delineate the unique biological signatures of IL-17F homodimers versus IL-17A/F heterodimers.

How do IL-17F and IL-17A differ in their contributions to inflammatory disease models?

IL-17F and IL-17A exhibit both overlapping and distinct roles in inflammatory disease models, which have been elucidated through studies with knockout mice and specific neutralizing antibodies:

• Asthma and Airway Inflammation:

  • IL-17A-deficient mice show reduced airway hyperresponsiveness in certain asthma models

  • IL-17F-deficient mice display different defects in experimental asthma models compared to IL-17A knockouts

  • Chronic overexpression of IL-17F in lung epithelium leads to lymphocyte and macrophage infiltration plus mucus hyperplasia, similar to IL-17A overexpression, but with delayed kinetics

  • Inhalation of recombinant IL-17F protein alone was insufficient to trigger neutrophil recruitment in some studies, unlike IL-17A

• Inflammatory Bowel Disease:

  • IL-17F and IL-17A knockout mice show distinct defects in colitis models

  • IL-17F may play either protective or pathogenic roles depending on the specific model and disease stage

  • The balance between IL-17F homodimers and IL-17A/F heterodimers may influence disease progression

• Autoimmune Diseases:

  • IL-17A has been more strongly implicated in rheumatoid arthritis and multiple sclerosis than IL-17F

  • Anti-IL-17A antibodies have shown remarkable clinical efficacy in psoriasis and psoriatic arthritis patients

  • The specific contribution of IL-17F to these conditions remains less well defined

• Host Defense:

  • Both IL-17A and IL-17F protect skin and mucosal barriers against infectious agents

  • IL-17F appears to have particularly important roles in mucosal immunity against extracellular bacteria and fungi

The biological potencies of IL-17A, IL-17A/F, and IL-17F in inducing inflammatory mediators correlate with their binding affinities toward IL-17RA, with IL-17A generally showing the highest potency and IL-17F the lowest .

What are the most promising therapeutic approaches targeting IL-17F for inflammatory diseases?

While IL-17A has been successfully targeted therapeutically with antibodies like secukinumab and ixekizumab for psoriasis and psoriatic arthritis, specific targeting of IL-17F or dual targeting of both IL-17A and IL-17F represents a promising frontier in cytokine-targeted therapy:

• Selective Anti-IL-17F Antibodies:

  • May provide benefit in diseases where IL-17F plays a predominant role over IL-17A

  • Could offer a more targeted approach with potentially fewer side effects

  • Would leave IL-17A-mediated host defense mechanisms intact

• Dual IL-17A/F Inhibitors:

  • Bimekizumab is a humanized monoclonal antibody that neutralizes both IL-17A and IL-17F

  • This approach may provide superior efficacy in conditions where both cytokines contribute to pathology

  • Would inhibit signaling from both homodimers and the IL-17A/F heterodimer

• Receptor-Targeted Approaches:

  • Blocking IL-17RA affects signaling by multiple IL-17 family members, potentially offering broader anti-inflammatory effects

  • Selective IL-17RC inhibitors might provide more focused inhibition of IL-17F and IL-17A responses

  • The discovery of potential IL-17RA-independent signaling through IL-17RC suggests that targeting both receptors might be necessary for complete inhibition

• Small Molecule Inhibitors:

  • Targeting the unique structural features of the IL-17F homodimer or its receptor binding interfaces

  • Developing inhibitors of downstream signaling components like Act1 or TRAF6

  • Creating small molecules that disrupt the formation of IL-17F homodimers

• Combination Therapies:

  • Combining IL-17F inhibition with targeting of related cytokines (e.g., IL-23, TNF-α)

  • Dual targeting of different inflammatory pathways may provide synergistic benefits

The choice of therapeutic approach should be guided by a deep understanding of the relative contributions of IL-17F versus IL-17A in specific disease contexts, and the potential consequences of inhibiting one or both cytokines for host defense.

How does genetic variation in IL-17F affect susceptibility to human diseases?

Genetic variation in IL-17F has been associated with susceptibility to several inflammatory and infectious diseases, providing insights into its role in human pathology:

• Single Nucleotide Polymorphisms (SNPs):

  • The IL-17F H161R polymorphism (rs763780), which results in a histidine to arginine substitution, has been linked to altered IL-17F function

  • This variant appears to act as a natural antagonist to wild-type IL-17F, potentially protecting against certain inflammatory conditions

  • Other SNPs in the IL-17F gene and its regulatory regions have been identified and may influence expression levels

• Disease Associations:

  • Inflammatory Bowel Disease: Certain IL-17F polymorphisms have been associated with susceptibility to ulcerative colitis and Crohn's disease in some populations

  • Asthma and Allergic Disorders: IL-17F gene variants have been linked to asthma risk and severity in several studies

  • Autoimmune Diseases: Associations with rheumatoid arthritis, multiple sclerosis, and systemic lupus erythematosus have been reported with varying degrees of consistency

  • Infectious Diseases: Some IL-17F polymorphisms may influence susceptibility to bacterial and fungal infections, particularly at mucosal surfaces

• Copy Number Variation:

  • The chromosomal region containing the IL-17F gene may be subject to copy number variations in some individuals

  • Such variations could potentially affect the balance between IL-17F and IL-17A expression

• Epigenetic Regulation:

  • Methylation patterns in the IL-17F promoter region have been associated with altered gene expression in certain disease states

  • Histone modifications affecting IL-17F expression may contribute to disease susceptibility

• Gene-Environment Interactions:

  • The impact of IL-17F genetic variations may depend on environmental factors like microbiome composition and pathogen exposure

  • Dietary factors and pollutants might interact with IL-17F genotypes to influence disease risk

Understanding the functional consequences of genetic variations in IL-17F can provide insights into personalized approaches to treating inflammatory diseases, potentially identifying patients who might benefit most from IL-17F-targeted therapies.

What are the most significant unresolved questions regarding IL-17F function and signaling?

Despite considerable advances in understanding IL-17F biology, several important questions remain unresolved:

• Signaling Mechanisms:

  • What is the precise role of the newly discovered symmetrical 2:1 complex between IL-17F and IL-17RC?

  • Do IL-17RA-independent signaling pathways exist for IL-17F, and if so, what are their biological consequences?

  • How do cells integrate signals from IL-17F homodimers versus IL-17A/F heterodimers?

• Heterodimer Biology:

  • What regulates the relative production of IL-17F homodimers versus IL-17A/F heterodimers in different cell types and conditions?

  • Does the IL-17A/F heterodimer have unique signaling properties beyond what would be expected from the combined actions of the homodimers?

  • How do the two faces of the IL-17A/F heterodimer contribute to receptor recruitment and signaling in living cells?

• Disease Relevance:

  • What is the precise role of IL-17F in human autoimmune and inflammatory diseases compared to IL-17A?

  • Are there diseases where IL-17F plays a dominant pathogenic role over IL-17A?

  • How does the balance between IL-17F, IL-17A, and IL-17A/F influence disease progression and treatment response?

• Cellular Sources and Regulation:

  • What regulates IL-17F expression in non-T cell sources such as innate lymphoid cells and myeloid cells?

  • How is IL-17F production regulated differently from IL-17A in various cellular contexts?

  • What epigenetic mechanisms control stable versus transient IL-17F expression?

• Therapeutic Targeting:

  • Will selective IL-17F inhibition provide therapeutic benefit in diseases not fully responsive to IL-17A blockade?

  • What are the consequences of long-term IL-17F inhibition for host defense and microbiome composition?

  • Can structure-based design yield small molecule modulators of IL-17F function with favorable therapeutic properties?

Addressing these questions will require interdisciplinary approaches combining structural biology, cell signaling analysis, immunology, and clinical investigation.

What novel experimental techniques might advance our understanding of IL-17F biology?

Several cutting-edge experimental approaches could significantly advance our understanding of IL-17F biology:

• Single-Cell Technologies:

  • Single-cell RNA sequencing to identify heterogeneity in IL-17F-producing cell populations

  • Mass cytometry (CyTOF) combined with IL-17F detection to characterize rare IL-17F-producing cells

  • Single-cell proteomics to analyze IL-17F-induced signaling at the individual cell level

• Advanced Imaging Techniques:

  • Super-resolution microscopy to visualize IL-17F receptor complex formation and trafficking

  • Live-cell FRET-based biosensors to monitor IL-17F-induced signaling in real-time

  • Intravital microscopy to track IL-17F-dependent cellular responses in vivo

• Structural Biology Innovations:

  • Cryo-electron microscopy to capture dynamic conformational changes in IL-17F-receptor complexes

  • Hydrogen-deuterium exchange mass spectrometry to map protein-protein interaction surfaces

  • AlphaFold and other AI-based prediction methods to model IL-17F interactions with novel binding partners

• Genetic Engineering Approaches:

  • CRISPR-based screens to identify novel components of IL-17F signaling pathways

  • Generation of reporter mice with fluorescent tags on endogenous IL-17F and IL-17A to visualize the dynamics of cytokine production

  • Creation of knock-in mice expressing obligate IL-17F homodimers or IL-17A/F heterodimers

• Systems Biology Approaches:

  • Multi-omics integration combining transcriptomics, proteomics, and metabolomics of IL-17F responses

  • Network analysis to identify key nodes in IL-17F signaling networks

  • Mathematical modeling of IL-17F signaling dynamics and competition with IL-17A

• Organoid and Tissue Engineering:

  • Epithelial organoids to study IL-17F effects on tissue barrier function

  • Microfluidic organ-on-chip models combining immune cells with tissue targets

  • 3D bioprinting to create complex tissue models for studying IL-17F in physiological contexts

These technological advances could help resolve the remaining questions about IL-17F biology and potentially reveal new therapeutic opportunities.

How might targeting the unique structural features of IL-17F lead to novel therapeutic approaches?

The unique structural features of IL-17F provide several opportunities for novel therapeutic development:

• Structure-Based Drug Design:

  • The crystal structure of IL-17F and its complexes with receptors provides templates for rational drug design

  • Small molecule inhibitors could be designed to target specific binding pockets at the cytokine-receptor interface

  • Peptide mimetics based on receptor binding epitopes could block IL-17F without affecting IL-17A

• Heterodimer-Specific Targeting:

  • The "two-faced" nature of the IL-17A/F heterodimer offers the possibility of developing antibodies or other biologics that specifically recognize the heterodimer interface

  • Such agents could selectively inhibit IL-17A/F without affecting IL-17A or IL-17F homodimers

  • This approach might provide more selective modulation of inflammatory responses

• Allosteric Modulators:

  • Since receptor binding induces allosteric changes in IL-17F, compounds that lock the cytokine in an inactive conformation could be effective inhibitors

  • Targeting allosteric sites distinct from the receptor binding interface might provide greater selectivity

  • Such modulators might alter signaling quality rather than completely blocking it

• Receptor-Specific Approaches:

  • The discovery that IL-17RC can form a symmetrical 2:1 complex with IL-17F suggests that targeting specific epitopes on IL-17RC might selectively modulate IL-17F signaling

  • Compounds that alter the stoichiometry of receptor complexes could fine-tune signaling outcomes

  • Bispecific antibodies targeting both IL-17RA and IL-17RC might have unique effects compared to blocking either receptor alone

• Signalosome Disruptors:

  • Targeting the protein-protein interactions involved in assembling the IL-17F signaling complex

  • Small molecules disrupting the interaction between IL-17F receptors and Act1 or TRAF6

  • Compounds that selectively inhibit specific downstream branches of IL-17F signaling

These structure-guided approaches could lead to a new generation of therapeutics with improved selectivity and potentially fewer side effects compared to current biologics targeting the IL-17 pathway.