IL17B Human, His

What is the basic structure of human IL-17B protein?

Human IL-17B is a homodimeric, non-disulfide-linked polypeptide that contains 180 amino acids per monomer with a molecular mass of approximately 20.4 kDa in monomeric form (36.5 kDa as a dimer). The recombinant form expressed in E. coli systems typically contains 322 amino acids (two chains of 161aa each) . Unlike other IL-17 family members which form disulfide-linked dimers, IL-17B exists as a non-covalently linked dimer. The protein contains a conserved cysteine-knot fold near the C-terminus, which is characteristic of the IL-17 family, but shows considerable sequence divergence at the N-terminus compared to other family members .

How does IL-17B differ from other IL-17 family members?

IL-17B shares only 29% homology with human IL-17A, making it structurally distinct despite belonging to the same cytokine family . While IL-17A, IL-17F, and IL-17C contribute primarily to neutrophil-mediated responses against bacteria and fungi, IL-17B (like IL-25) is functionally associated with type 2 immune responses . A critical distinction is that IL-17B is never detected in activated CD4+ T cells (the primary source of IL-17A) . Additionally, IL-17B has a different tissue expression pattern compared to other family members, with strong expression in pancreas, small intestine, stomach, and testis .

What are the optimal storage and reconstitution conditions for recombinant IL-17B?

For long-term storage, lyophilized IL-17B should be kept desiccated below -18°C, where it remains stable for at least 12 months from the date of receipt . While the lyophilized protein can remain stable at room temperature for up to 3 weeks, this is not recommended for valuable research material. Upon reconstitution, IL-17B should be stored at 4°C if used within 2-7 days. For future use, store reconstituted protein below -18°C, preferably with a carrier protein (0.1% HSA or BSA) to prevent adhesion to surfaces and improve stability . Reconstitution should be performed in sterile water to a concentration not less than 100μg/ml, which can then be further diluted to other aqueous solutions as needed for experimental applications .

What detection methods are most effective for studying IL-17B expression and activity?

Multiple complementary approaches should be employed for comprehensive IL-17B detection:

• Western blotting: Anti-human IL-17B antibodies (such as clone 174113) can effectively detect IL-17B in cell lysates, though researchers should be aware of potential cross-reactivity with mouse IL-17B (approximately 25%) .

• Flow cytometry: For intracellular detection, cells should be fixed with paraformaldehyde and permeabilized with saponin before staining with specific monoclonal antibodies. This approach has been validated in K562 human chronic myelogenous leukemia cells .

• ELISA: Sandwich immunoassay techniques allow for quantitative measurement of IL-17B in supernatants or serum samples.

• RT-PCR and qPCR: These techniques can detect IL-17B mRNA expression, particularly useful when analyzing tissue-specific expression patterns.

When selecting detection methods, consider that IL-17B expression varies significantly between tissues, with highest levels in pancreas, small intestine, stomach, and testis, while being weakly expressed in spinal cord, prostate, colon, and ovary .

How can I design experiments to study IL-17B signaling pathways?

To effectively study IL-17B signaling pathways, consider these methodological approaches:

• Receptor binding studies: Since IL-17B binds to IL-17RB with approximately 30-fold lower affinity than IL-25, binding kinetics experiments should employ a wide concentration range. Note that IL-17B has a similar association rate (Kon) but substantially faster dissociation rate (Koff) compared to IL-25 .

• Downstream signaling: Include analysis of TRAF6 recruitment, as IL-17RB contains a unique TRAF6-binding domain not found in IL-17RA . Monitor activation of NF-κB and MAPK pathways.

• Cell models: Select appropriate cell types expressing IL-17RB, such as innate type 2 lymphocytes, NKT cells, and Th2 cells for immune function studies . For studying cancer-related functions, breast cancer cell lines with HER2 amplification have shown significant IL-17B responsiveness .

• Receptor blocking: Design experiments using neutralizing antibodies against both IL-17RA and IL-17RB, as functional studies indicate IL-17B uses both receptors for signaling .

• Cytokine production: Measure type 2 cytokine secretion (IL-4, IL-5, IL-13) as readouts for IL-17B activity, particularly in experiments examining IL-17B's ability to augment IL-33-driven responses .

What are the challenges in producing functional recombinant IL-17B?

Producing functional recombinant IL-17B presents several technical challenges:

• Proper dimerization: Unlike other IL-17 family members that form disulfide-linked dimers, IL-17B forms non-covalent dimers, making proper folding and assembly more challenging to achieve and maintain .

• Expression systems: E. coli-based expression systems are commonly used, but researchers should verify that the protein maintains its native conformation and biological activity through functional assays .

• Glycosylation: Native IL-17B is a glycoprotein, but E. coli-produced recombinant versions lack glycosylation, which may affect certain aspects of protein function or stability .

• Solubility and aggregation: Maintaining protein solubility requires careful buffer optimization; recommendations include reconstitution in sterile water at concentrations not less than 100μg/ml before dilution in application-specific buffers .

• Activity verification: Functional testing is essential, as improperly folded protein may retain immunoreactivity for detection assays but lack biological activity. Testing should include receptor binding assays and cell-based functional assays measuring appropriate cytokine responses.

How does the IL-17B signaling mechanism differ from that of IL-17A?

IL-17B signaling exhibits several distinct differences from IL-17A signaling:

• Receptor complex: IL-17B signals through both IL-17RA and IL-17RB receptor subunits, forming a heterodimeric complex similar to IL-25, whereas IL-17A signals through IL-17RA/IL-17RC heterodimers .

• Receptor binding domains: While both cytokines utilize IL-17RA, IL-17B interacts primarily with IL-17RB which contains a unique TRAF6-binding domain not present in IL-17RA .

• Downstream effects: IL-17A predominantly induces pro-inflammatory cytokines (TNF-α, IL-6) and chemokines (CXCL8, CXCL1) involved in neutrophil recruitment and activation, whereas IL-17B promotes type 2 immune responses characterized by production of IL-4, IL-5, and IL-13 .

• Cellular sources: IL-17A is primarily produced by Th17 CD4+ T cells, while IL-17B has never been detected in these cells. Instead, IL-17B is produced by neutrophils, chondrocytes, neurons, and various B cell subsets .

• Target cells: IL-17B acts on innate type 2 lymphocytes, NKT cells, and CD4+ CRTH2+ Th2 cells, having a functional profile more similar to IL-25 than to IL-17A .

What is the relationship between IL-17B and type 2 immune responses?

IL-17B functions as a regulator of type 2 immunity with several key characteristics:

• Target cell populations: IL-17B elicits type 2 cytokine secretion from innate type 2 lymphocytes, NKT cells, and CD4+ CRTH2+ Th2 cells, similar to IL-25 but distinct from IL-17A .

• Receptor dependency: This activity depends on both IL-17RA and IL-17RB receptor subunits, with signaling through this heterodimeric complex being essential for downstream effects .

• Synergistic activity: IL-17B can augment IL-33-driven type 2 responses, indicating potential crosstalk between these pathways in allergic or parasite-induced immune responses .

• Physiological context: These functions position IL-17B as a novel component in the regulation of human type 2 immunity, potentially contributing to antiparasitic defense mechanisms but also potentially involved in allergic responses when dysregulated .

The similarity in function between IL-17B and IL-25, despite their limited sequence homology, suggests evolutionary conservation of type 2 response-inducing activity within the IL-17 family, possibly representing functional redundancy or context-specific regulation .

How can researchers differentiate between IL-17B and IL-25 (IL-17E) effects in experimental systems?

Differentiating between IL-17B and IL-25 effects requires careful experimental design:

• Binding affinity considerations: IL-17B binds to IL-17RB with approximately 30-fold lower affinity than IL-25, necessitating different concentration ranges in experimental settings. Dose-response experiments should be performed to determine threshold concentrations for each cytokine .

• Receptor blocking strategies:

  • Use anti-IL-17RB antibodies to block both cytokines

  • Use specific neutralizing antibodies against each cytokine

  • Design receptor mutants with differential binding to IL-17B versus IL-25

• Expression pattern analysis: Examine natural expression patterns, as IL-17B and IL-25 may be produced by different cell types or in response to different stimuli in your model system .

• Gene silencing approaches: Employ siRNA or CRISPR-based gene targeting specifically against either IL-17B or IL-25 to determine their individual contributions.

• Kinetic studies: Analyze the kinetics of responses, as IL-17B has a faster dissociation rate (Koff) from IL-17RB compared to IL-25, potentially resulting in differences in signaling duration or intensity .

What is the evidence for IL-17B involvement in cancer progression?

Multiple lines of evidence support IL-17B's role in cancer progression:

• Prognostic correlations: High expression of IL-17B or its receptor IL-17RB has been associated with poor prognosis in multiple cancer types, including breast cancer. In a cohort of 143 breast cancer patients, both IL-17B and IL-17RB expression correlated with reduced patient survival .

• Receptor upregulation: IL-17RB was found to be upregulated in 19% of patients with ductal invasive breast carcinoma, with its detection significantly correlated with poor prognosis and high mortality rates .

• HER2 association: In breast cancer, IL-17RB expression is associated with HER2 amplification, with the lowest survival rates observed in patients with high expression of both IL-17RB and HER2 .

• Functional mechanisms: IL-17B signaling through IL-17RB directly promotes cancer cell survival, proliferation, and migration, and can induce resistance to conventional chemotherapeutic agents .

• Molecular pathway analysis: Analysis of microarray data from 1809 breast cancer patients showed high IL-17B expression was significantly correlated with poorer prognosis in the whole population and particularly in basal-like breast cancer subtypes .

These findings collectively suggest the IL-17B/IL-17RB signaling axis as a potential therapeutic target in cancer, particularly in breast cancer where expression correlates with aggressive disease features.

How does IL-17B contribute to inflammatory and autoimmune conditions?

IL-17B has complex roles in inflammatory and autoimmune conditions:

• Rheumatoid arthritis: IL-17B expression has been detected in rheumatoid synovial tissues from patients with rheumatic arthritis, where it is mainly produced by neutrophils and may contribute to joint inflammation .

• Inflammatory bowel disease: IL-17B was originally described as increased during intestinal inflammation, potentially contributing to disease pathogenesis through stimulation of TNF-α and IL-1β production by monocytic cells .

• Pro-inflammatory effects: IL-17B promotes neutrophil migration upon intraperitoneal administration, suggesting a direct pro-inflammatory role in certain contexts .

• Type 2 inflammation: Through its ability to elicit type 2 cytokine production and augment IL-33-driven responses, IL-17B may contribute to type 2 inflammatory conditions such as asthma and atopic dermatitis, where IL-17RB expression is elevated .

• Protective inflammation: Physiologically, IL-17B-mediated inflammation may contribute to host defense against extracellular pathogens, particularly in mucosal tissues where it is expressed .

The precise role of IL-17B in different inflammatory conditions remains to be fully elucidated, with evidence suggesting both pathogenic and protective roles depending on the specific disease context and tissue environment.

What are the physiological roles of IL-17B in healthy tissues?

Despite being primarily studied in disease contexts, IL-17B has several important physiological functions:

• Host defense: IL-17B contributes to protection against extracellular pathogens, particularly at mucosal surfaces where it is expressed. This protective function operates through mechanisms distinct from IL-17A-mediated defense .

• Epithelial integrity: Along with other IL-17 family members, IL-17B helps maintain epithelial barrier integrity, which is crucial for preventing invasion by microorganisms and environmental toxins .

• Metabolic regulation: IL-17B may modulate adipocyte activity, suggesting potential roles in energy metabolism and adipose tissue homeostasis .

• Neuronal functions: IL-17B is expressed in neurons and may have roles in regulating cognitive processes, though this area requires further investigation .

• Immune homeostasis: Through its ability to regulate type 2 immune responses, IL-17B likely contributes to normal immune homeostasis, particularly in tissues where it is highly expressed such as the gastrointestinal tract .

Understanding these physiological roles is crucial when targeting the IL-17B pathway therapeutically, as complete blockade could potentially disrupt important homeostatic functions.

How might targeting IL-17B differ from current IL-17A-targeted therapies?

Therapeutic targeting of IL-17B would have several distinct implications compared to current IL-17A-targeted approaches:

• Immune response profile: While IL-17A antagonists (secukinumab, ixekizumab) primarily affect neutrophil-mediated immunity and Th17 responses, IL-17B targeting would impact type 2 immune responses, potentially affecting conditions like asthma or parasitic infections differently .

• Side effect profile: IL-17A blockade increases risk of fungal and bacterial infections and can exacerbate inflammatory bowel disease in some patients. IL-17B antagonism would likely produce a different pattern of side effects based on its physiological roles in epithelial integrity and type 2 immunity .

• Cancer implications: Given IL-17B's association with cancer progression and poor prognosis, targeting IL-17B might have anti-cancer effects not observed with IL-17A antagonists, particularly in breast cancers with HER2 amplification .

• Receptor targeting strategy: Since IL-17B signals through IL-17RA/IL-17RB heterodimers, receptor-targeted approaches must account for potential effects on IL-25 signaling, which uses the same receptor complex. Selective targeting of IL-17B over IL-25 would require sophisticated drug design strategies .

• Dual targeting potential: Combined blockade of IL-17A and IL-17B pathways might offer synergistic benefits in conditions with mixed inflammatory profiles, addressing both neutrophilic and type 2 inflammatory components simultaneously.

What methodological approaches are needed to resolve conflicting reports about IL-17B functions?

Resolving contradictory findings regarding IL-17B functions requires rigorous methodological approaches:

• Species-specific analysis: Clearly differentiate between human and mouse IL-17B functions, as there may be significant species differences. Human IL-17B shares only 88% homology with its murine ortholog, potentially resulting in functional differences .

• Context-dependent studies: Systematically evaluate IL-17B functions in different tissue and disease contexts, as effects may vary significantly based on the microenvironment and presence of other cytokines.

• Standardized recombinant proteins: Use well-characterized, functional recombinant proteins with demonstrated bioactivity. Poor protein quality or inadequate functional validation may contribute to conflicting results.

• Receptor complex analysis: Employ techniques to definitively establish receptor usage in each experimental system, as IL-17B may signal through different receptor complexes in different contexts .

• Comprehensive readouts: Measure multiple outcome parameters rather than focusing on single readouts, as IL-17B may have pleiotropic effects that are missed when examining limited endpoints.

• Genetic validation: Complement in vitro studies with genetic approaches (knockout/knockin models) to verify physiological relevance of observed effects.

• Single-cell analysis: Apply single-cell technologies to identify specific cellular sources and targets of IL-17B, which may help explain apparently contradictory functions.

What are the potential applications of IL-17B in biomarker development for cancer prognostication?

IL-17B shows significant promise as a biomarker in several cancer contexts:

• Prognostic stratification: Given the correlation between IL-17B/IL-17RB expression and poor outcomes in breast cancer, developing standardized assays to quantify expression levels could help identify high-risk patients who might benefit from more aggressive treatment approaches .

• Subtype classification: In breast cancer, IL-17RB expression associates with HER2 amplification, suggesting potential utility in molecular subtyping or refinement of existing classification systems .

• Therapeutic response prediction: Evaluating IL-17B pathway activation might predict response to certain therapies, particularly in cancers where this pathway promotes chemoresistance.

• Multi-marker panels: Integration of IL-17B with other biomarkers could improve prognostic accuracy compared to single markers alone. Particularly valuable would be combining IL-17B with established markers like HER2, where synergistic prognostic value has been observed .

• Liquid biopsy development: Investigating whether circulating IL-17B levels in blood correlate with tissue expression could enable non-invasive monitoring of this pathway in cancer patients.

Implementation would require standardized detection methods with established cutoff values for clinical decision-making, validation in large prospective cohorts, and demonstration of added value beyond conventional prognostic factors.

What are the key unanswered questions about IL-17B that warrant further investigation?

Several critical knowledge gaps remain in our understanding of IL-17B:

• Transcriptional regulation: What factors control IL-17B expression in different tissues and disease states? This fundamental question remains poorly characterized compared to other IL-17 family members.

• Receptor complex dynamics: How do IL-17RA and IL-17RB interact to form functional signaling complexes, and what determines preferential binding of IL-17B versus IL-25 in physiological settings?

• Signaling pathway specificity: What downstream signaling components are uniquely activated by IL-17B compared to other family members, and how does this translate to distinct functional outcomes?

• Evolutionary significance: Why has IL-17B function been conserved evolutionarily when IL-25 appears to serve similar functions in type 2 immunity? Does IL-17B play unique roles in specific contexts?

• Cancer-promoting mechanisms: Through what precise molecular mechanisms does IL-17B promote cancer progression, chemoresistance, and metastasis?

• Therapeutic potential: Could targeting IL-17B prove beneficial in cancer or inflammatory conditions, and how might this compare to existing therapies targeting related pathways?

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

How might advances in single-cell technologies enhance our understanding of IL-17B biology?

Single-cell technologies offer transformative potential for IL-17B research:

• Source identification: Single-cell RNA sequencing (scRNA-seq) can precisely identify cellular sources of IL-17B in different tissues and disease states, resolving current controversies about expression patterns .

• Receptor co-expression: Single-cell analysis can map cells co-expressing IL-17RA and IL-17RB, identifying potential target populations for IL-17B action that may have been missed in bulk tissue analyses.

• Response heterogeneity: Single-cell approaches can reveal the heterogeneity in cellular responses to IL-17B stimulation, potentially identifying previously unrecognized responsive cell populations.

• Pathway reconstruction: Combining single-cell transcriptomics with computational approaches allows reconstruction of signaling pathways activated by IL-17B in specific cell types.

• Spatial context: Spatial transcriptomics can place IL-17B-producing and responding cells in their tissue context, revealing microanatomical relationships that influence function.

• Temporal dynamics: Single-cell trajectory analysis can track how IL-17B signaling evolves over time in developmental or disease processes.

Implementation of these approaches could resolve long-standing questions about IL-17B function and potentially identify novel therapeutic opportunities targeting specific cellular interactions.