IFNL4 Antibody

What is IFNL4 and how does it function in viral immunity?

IFNL4 (Interferon Lambda 4) is a noncanonical interferon with unusual functional characteristics. The IFNL4 gene encodes the IFN-λ4 protein which demonstrates antiviral activity in vitro . Paradoxically, genetic evidence reveals that patients with a nonfunctional IFNL4 gene exhibit increased clearance of hepatitis C virus (HCV) but also increased liver inflammation, suggesting that IFN-λ4 has proviral and anti-inflammatory effects in vivo . This unique behavior appears to be related to its distinctive transcriptional regulation, as IFNL4 is poorly induced by viral infections compared to canonical interferons like IFNB1 and IFNL1 .

In humoral immunity research, studies indicate that the ability to produce IFN-λ4 augments antibody levels against HCV, suggesting association with T helper cell 2 (Th2) immune skewing . Additionally, intracellular accumulation of IFN-λ4 can induce endoplasmic reticulum (ER) stress, potentially contributing to antiproliferative mechanisms with implications for HCV progression, cirrhosis, and hepatocellular carcinoma development .

What genetic polymorphisms affect IFNL4 expression and function?

Several key genetic polymorphisms determine IFNL4 expression and functionality:

• The rs368234815 (ΔG/TT) polymorphism dictates whether the IFNL4 gene produces the IFN-λ4 protein. The ΔG allele permits protein production, while the TT allele creates a frameshift preventing protein expression .

• The rs12979860 polymorphism, in strong linkage disequilibrium with rs368234815, influences susceptibility to chronic HCV infection. Homozygous C allele carriers (CC genotype) demonstrate increased HCV clearance compared to CT or TT genotypes .

• The rs117648444 (G>A) polymorphism affects IFN-λ4 protein functionality through a proline to serine substitution at position 70 (P70S), creating four possible haplotypes :

  • TT/G or TT/A: IFNλ4-Null (no protein production)

  • ΔG/G: IFNλ4-P70 (fully active protein)

  • ΔG/A: IFNλ4-S70 (impaired activity)

Patients with IFNλ4-Null and IFNλ4-S70 variants exhibit lower hepatic interferon-stimulated gene expression, associated with increased viral clearance and improved response to IFN-based therapy compared to patients with the IFNλ4-P70 variant .

How does IFNL4 differ from other interferons in expression and regulation?

IFNL4 demonstrates markedly different expression patterns compared to canonical interferons like IFNB1 and IFNL1. Experimental studies show that IFNL4 is poorly induced by viral infections, with expression levels much lower than other interferons even after viral challenge . Quantitatively, the absolute expression of IFNB1 and IFNL1 was approximately 1,000-fold higher than IFNL4 following viral infection in most cell types tested .

Promoter studies reveal that the IFNL1 promoter generates approximately 230-fold stronger luciferase activity than the IFNL4 promoter . Furthermore, while the IFNL1 promoter is potently induced by Sendai virus infection, the IFNL4 promoter shows minimal response . This indicates that while the IFNL4 promoter is highly conserved and functional, it lacks the defining characteristic of interferons: effective induction by viral infections .

How do IFNL4 genotypes correlate with antibody responses in HCV infection?

Research demonstrates significant correlation between IFNL4 genotypes and antibody responses in HCV infection. Patients producing IFN-λ4 (those with IFNL4 rs12979860 CT/TT genotypes) exhibit higher signal to cut-off (S/CO) anti-HCV antibody ratios compared to non-producers (IFNL4 rs12979860 CC genotype) . This association remains significant in multivariate analysis, indicating that IFN-λ4 production independently enhances antibody levels against HCV .

This finding suggests IFN-λ4 may promote T helper cell 2 (Th2) immune skewing, with potential clinical implications beyond HCV infection . Recent research identified an association between the IFNL4 haplotype related to IFNλ4-P70 production and lower antibody response levels , indicating a complex relationship between IFNL4 variants and humoral immunity.

Additional factors independently associated with higher anti-HCV antibody levels include male gender, absent/mild steatosis, and lower viral load .

How can researchers reconcile IFNL4's paradoxical effects in vitro versus in vivo?

The paradoxical behavior of IFNL4—antiviral in vitro but apparently proviral in vivo—requires sophisticated experimental approaches:

• Integrated model systems: Develop experimental designs that bridge cellular and clinical observations using primary human hepatocytes in 3D culture systems or humanized mouse models that better approximate human physiology .

• Context-dependent analyses: Since IFNL4 functions differently depending on context, experiments should incorporate multiple cell types, varying viral loads, and different inflammatory conditions to capture the full spectrum of responses .

• Temporal dynamics investigation: Design time-course studies that monitor effects across multiple timepoints, from acute to chronic infection phases .

• Combined genomic-functional approaches: Integrate genetic analyses (IFNL4 genotyping) with functional assays (measuring ISG expression, viral replication, immune responses) to correlate genotype with phenotype .

• Systems biology integration: Employ computational modeling to integrate data from genomic, transcriptomic, proteomic, and cellular levels to develop comprehensive models of IFNL4 function .

• Intracellular trafficking assessment: Given evidence of IFN-λ4 intracellular accumulation inducing ER stress, experimental designs should carefully assess protein localization, trafficking, and associated stress responses .

Researchers must acknowledge model limitations, particularly the absence of IFNL4 in mouse genomes, which complicates animal studies .

What mechanisms explain IFN-λ4's contradictory antiviral activity in vitro versus proviral effects in vivo?

Several mechanisms potentially explain IFNL4's paradoxical effects:

How do host IFNL4 variants interact with viral genetic variations to modulate antibody responses?

Research investigating host-pathogen genetic interactions reveals a complex interplay between IFNL4 variants and viral polymorphisms in determining antibody responses:

• Dual genetic determinants: Both host genetic factors (IFNL4 variants) and viral genetic factors (HCV E1/E2 coding region polymorphisms) independently and jointly influence antibody responses .

• Viral epitope variations: In HCV genotype 3a infection, three amino acid substitutions in the HCV E2 protein associate with anti-E1/E2 antibody binding and neutralization sensitivity. Additionally, variants in two N-linked glycosylation sites correlate with antibody responses .

• IFNL4 haplotype effects: The IFNL4 haplotype producing IFNλ4-P70 associates with lower antibody response levels, suggesting a specific mechanism by which host genetics affects humoral immunity .

The methodological approach employed by researchers included:

• Genetic screening to identify viral polymorphisms linked to antibody responses

• Binding and neutralization assays using HCV pseudotype-based infection models

• Linear regression analysis to determine associations between genetic variants and antibody function

Researchers controlled for sex, cirrhosis status, prior interferon treatment, age, and viral load when assessing genetic associations. Males had lower anti-E1E2 binding levels than females, and patients without cirrhosis or with previous interferon treatment displayed lower antibody binding responses .

What methodological considerations apply when studying IFNL4 in animal models given its absence in mice?

The absence of IFNL4 in mouse genomes presents significant challenges for researchers . Several methodological approaches can address this limitation:

• Non-murine models: Consider alternative animal models that express IFNL4. The functional IFNL4 gene is conserved in all other primates , suggesting non-human primates may serve as more appropriate models.

• Transgenic approaches: Develop transgenic mice expressing human IFNL4 variants (P70, S70) under appropriate regulatory control to investigate its function in vivo.

• Humanized systems: Employ humanized liver mouse models where human hepatocytes are engrafted into immunodeficient mice, enabling study of IFNL4 in a more relevant cellular context for hepatotropic viruses.

• Advanced human cell systems: Given animal model limitations, sophisticated human cellular models become especially important:

  • Primary human hepatocytes

  • Patient-derived organoids

  • Immune cell co-culture systems

  • 3D culture systems that better recapitulate tissue architecture

• Evolutionary biology perspective: Study the evolutionary loss of IFNL4 function in mice compared to its conservation in primates to gain insights into its biological significance.

• Genetic manipulation: The CRISPR/Cas9 approach used to create IFNLR1 knockout cells suggests similar techniques could investigate IFNL4 signaling components.

Researchers must acknowledge species differences and carefully consider how these might affect result interpretation when using any of these approaches.