The results of a recent study published in the Journal of Investigative Dermatology could have significant implications for the interpretation of IL36RN variants in diagnostic settings, potentially leading to improved management and treatment strategies for patients with generalized pustular psoriasis (GPP).
In addition, the study provides new insights into the structural elements that govern IL-36Ra activity, offering potential pathways for designing novel IL-36 inhibitors for the treatment of skin inflammation.
“We experimentally characterized the effects of 30 IL36RN variants on protein stability,” the researchers explained. “In parallel, we used a machine-learning tool (Rhapsody) to analyse the IL-36Ra three-dimensional structure and predict the impact of all possible amino acid substitutions.”
The researchers analyzed 30 variants distributed along the entire protein sequence, including 17 rare changes observed in patients with GPP (patient variants).
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By using mutagenized constructs for the 30 selected changes, the researchers confirmed that p.Pro27Leu and p.Ser113Leu, already known as destabilizing mutations, were associated with a significant reduction in the levels of IL-36Ra levels. In addition, 3 other GPP alleles (p.Ile42Asn, p.Glu112Lys, and p.Thr123Arg) were found to display a similar effect.
All destabilising changes were associated with high (greater than 50%) likelihoods of pathogenicity, according to machine learning analysis.
In contrast to population variants, most patient alleles led to the upregulation of IL-36 signaling. The results of computational alanine scanning showed that most (91%) residue changes had a mild or moderate destabilizing effect on the IL-36R/IL-36Ra interaction. Altogether, the researchers identified 13 amino acids that were critical to the IL-36Ra/IL36R interaction.
Moreover, the results of the in silico saturation mutagenesis experiments, which aimed to simulate all possible amino acid substitutions for the 155 residues in the IL-36Ra protein, indicated that the 15 most intolerant residues to mutations were more likely to be located within β-strands and the protein core.
These residues were affected by specific mutations (p.His32Arg, p.Arg48Trp, p.Pro76Leu, p.Arg102Trp, and p.Glu112Lys) and were more frequently found on the IL-36Ra/IL-36R binding interface than the remaining IL-36Ra amino acids.
Reference
Hassi NK, Weston T, Rinaldi G, et al. In-silico and in-vitro analysis of IL36RN mutations reveals critical residues for the function of the interleukin-36 receptor complex. J Invest Dermatol. Published online July 4, 2023. doi:10.1016/j.jid.2023.06.191
