IL 29 Human, His

What is IL-29 and what are its alternative designations in scientific literature?

IL-29 is a cytokine belonging to the type III interferons group, also termed interferons λ (IFN-λ). It plays an important role in immune responses against pathogens, particularly viruses, by mechanisms similar to type I interferons but targeting primarily cells of epithelial origin and hepatocytes. IL-29 is alternatively designated as interferon lambda 1 (IFN-λ1) and belongs to the larger IL-10 family of cytokines, which includes IL-10, IL-19, IL-20, IL-22, IL-24, IL-26, IL-28A, and IL-28B .

What is the structure and receptor binding mechanism of IL-29?

IL-29 is structurally related to the IL-10 family, but its primary amino acid sequence and function are more similar to type I interferons. The protein binds to a heterodimeric receptor complex consisting of:

• A specific subunit (IFNL1R, also called IL-28R1)

• A second subunit (IL-10R2) that is shared among the IL-10 family cytokines

This receptor binding initiates downstream signaling cascades that lead to antiviral and immunomodulatory effects. The structural characteristics of IL-29 enable it to function as an intermediate between the IL-10 family and type I interferons, giving it unique biological properties .

How is IL-29 encoded in the human genome?

IL-29 is encoded by the IFNL1 gene located on the long arm of chromosome 19 in humans. The gene consists of 5 exons. Interestingly, IL-29 is a pseudogene in mice, meaning the IL-29 protein is not produced in them . This has important implications for research, as mouse models cannot be used to directly study the function of IL-29, requiring alternative approaches such as humanized mice or other animal models.

What are the primary signaling pathways activated by IL-29?

IL-29 binding to its receptor activates multiple signaling pathways:

• JAK-STAT pathway - IL-29 signals predominantly through Janus kinase/signal transducers and activators of transcription (JAK-STAT), activating STAT1, STAT2, STAT3, and STAT5 .

• NF-κB pathway - IL-29 induces nuclear factor kappa B (NF-κB) signaling, contributing to inflammatory responses .

• MAPK pathway - IL-29 activates mitogen-activated protein kinase (MAPK), which is potentially involved in autoimmune disease development .

These signaling cascades lead to the expression of interferon-stimulated genes (ISGs) and the production of antiviral proteins, as well as modulation of immune cell functions .

How does histidine-tagged recombinant IL-29 differ from native IL-29 in experimental applications?

Histidine-tagged (His-tagged) recombinant IL-29 contains a string of histidine residues added during recombinant expression to facilitate protein purification through metal affinity chromatography. Key considerations for researchers include:

• Biological activity: His-tagged IL-29 generally retains functional activity comparable to native IL-29, but minor differences in receptor binding kinetics may occur.

• Expression systems: Both E. coli and mammalian cell (e.g., NS0) expression systems have been used to produce His-tagged IL-29, with the E. coli-derived version being established as the WHO international reference reagent .

• Structural considerations: The His-tag might influence protein folding or complex formation in some experimental contexts, requiring validation in specific applications.

• Standardization: When using His-tagged IL-29, researchers should calibrate against the WHO reference reagent (10/176) to ensure reproducibility across studies .

What are the optimal bioassay methods for measuring IL-29 activity?

Several validated bioassay methods can be used to measure IL-29 activity, each with specific advantages:

Antiviral assays:

• Measure IL-29-induced reduction of cytopathic effect in human cell lines challenged with viruses (e.g., encephalomyocarditis virus or vesicular stomatitis virus)

• Quantify protection using vital stain absorption

Reporter gene assays:

• HEK 293 cells transfected with secreted alkaline phosphatase cDNA linked to the ISRE promoter

• HuH7 cells transfected with an IFN-regulated firefly luciferase construct

STAT activation assays:

• Phospho-tyrosine STAT-1 ELISA methods using cell lysates after IL-29 stimulation

• Electrophoretic mobility shift assay (EMSA) to assess STAT-1 activation

How does IL-29 compare functionally with type I interferons in antiviral responses?

While IL-29 and type I interferons (e.g., IFN-α) share functional similarities, they have important distinctions:

Similarities:

• Both induce JAK-STAT signaling pathways

• Both upregulate interferon-stimulated genes including MxA, 2-5A synthetase, and class I MHC antigen

• Both possess antiviral properties

Key differences:

• Receptor specificity: IL-29 binds to IFNL1R/IL10R2 heterodimer, while type I IFNs bind to IFNAR1/IFNAR2

• Tissue distribution: IL-29 receptors are primarily expressed on epithelial cells and hepatocytes, giving it a more restricted target range than type I IFNs

• Clinical profile: Pegylated IL-29 shows fewer undesirable side effects compared to pegylated IFN-α2 in hepatitis C treatment

These differences make IL-29 a potentially valuable alternative to type I interferons in therapeutic applications, particularly for hepatitis C treatment, as demonstrated in the "EMERGE" phase IIb trial which compared pegylated IL-29 with pegylated IFN-α2a .

What is known about the role of IL-29 in autoimmune diseases?

IL-29 has been implicated in various autoimmune diseases:

Graves' Orbitopathy (GO):

• Higher levels of IL-29 in euthyroid patients with Graves' disease and active GO compared to those without GO

• IL-29 concentration: 165 (133-747) vs. 62 (62-558) pg/mL, p = 0.031

• ROC analysis identified an IL-29 cut-off of 105 pg/mL (sensitivity 1.000, specificity 0.597) as significantly indicating GO presence

Other autoimmune conditions with elevated IL-29 levels:

• Sjögren syndrome

• Rheumatoid arthritis

• Systemic sclerosis

• Systemic lupus erythematosus

• Psoriasis

• Hashimoto's thyroiditis

The involvement of IL-29 in these conditions appears related to its effects on JAK-STAT, MAPK, and NF-κB signaling pathways. IL-29 can influence B lymphocyte function and stimulate monocytes to secrete IL-6, IL-8, and IL-10, which may contribute to autoimmune pathogenesis .

What methodological considerations are important when studying IL-29 in immunological research?

When designing experiments to investigate IL-29 function in immunological research, several methodological considerations are crucial:

Cell type selection:

• Focus on cells expressing the IL-29 receptor complex (epithelial cells, hepatocytes)

• Consider plasmacytoid dendritic cells (pDCs), which show reduced expression of IFN-γ, IL-13, and IL-10 when treated with IL-29

Receptor expression analysis:

• Verify expression of both receptor components (IFNL1R and IL10R2) in your experimental system

• Consider receptor regulation in different contexts (e.g., inflammation, viral infection)

Standardization protocols:

• Use the WHO international reference reagent for IL-29 (preparation 10/176, E. coli-derived) with assigned unitage of 5,000 reference units per ampoule

• Include appropriate controls to account for inter-assay variability

Functional readouts:

• Monitor surface marker expression changes (e.g., CD80, CD83, ICOS-L, CCR7, CD62L on pDCs)

• Assess IL-29's effects in combination with other cytokines (e.g., IL-29 with IFN-α enhances costimulatory molecule expression)

How can researchers differentiate between the effects of IL-29 and other members of the IL-10 family?

Differentiating the specific effects of IL-29 from other IL-10 family members requires targeted experimental approaches:

Receptor targeting:

• Use blocking antibodies specifically targeting IL-29's unique receptor component (IFNL1R) versus the shared component (IL10R2)

• Develop receptor knockout cell lines using CRISPR/Cas9 to selectively eliminate IL-29 signaling

Specificity controls:

• Include multiple IL-10 family members in parallel experiments

• Use cytokine-specific neutralizing antibodies to confirm specific IL-29 effects

• Perform receptor expression profiling in your experimental system

Functional assays:

• Analyze transcriptional profiles induced by different IL-10 family members

• Compare effects on specific cell types where differential receptor expression occurs

• Examine downstream signaling pathway activation patterns unique to IL-29

What is the current status of IL-29 in therapeutic development?

IL-29 has shown promise as a therapeutic agent in several contexts:

Chronic Hepatitis C Infection:

• Pegylated IL-29 (IFN-λ1) has been evaluated in clinical trials as an alternative to pegylated IFN-α2

• Phase Ib trials demonstrated the potential of pegylated IL-29 with or without ribavirin

• The "EMERGE" phase IIb trial compared pegylated IL-29 with pegylated IFN-α2a in treatment-naïve patients with HCV genotypes 1 or 4

• Results showed similar sustained virological response rates between the two regimens but fewer side effects with IL-29

Potential advantages of IL-29 therapy:

• More targeted effects on epithelial cells and hepatocytes

• Reduced systemic side effects compared to type I interferons

• Good safety profile in chronic HCV-infected patients

How should IL-29 activity be standardized for research and therapeutic applications?

Standardization of IL-29 activity is critical for consistent research and therapeutic development:

WHO reference standard:

• In 2012, the WHO established preparation 10/176 (E. coli-derived IL-29) as the international reference reagent

• This standard has an assigned unitage of 5,000 reference units per ampoule

Standardization methodology:

• International collaborative studies have assessed different IL-29 preparations in various bioassays

• Both NS0-derived (07/212) and E. coli-derived (10/176) preparations were evaluated, with the latter being chosen as the reference standard

Experimental considerations:

• Researchers should calibrate in-house standards against the WHO reference

• Report IL-29 activity in reference units rather than arbitrary units

• Specify the expression system used (E. coli vs. mammalian cells)

• Document the specific bioassay used for potency determination