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Case Reports
. 2025 Nov;108(5):576-581.
doi: 10.1111/cge.14763. Epub 2025 Apr 30.

A Novel Intronic Variant in the KH3 Domain of HNRNPK Leads to a Mild Form of Au-Kline Syndrome

Maura Mingoia  1 , Alessandra Meloni  1 , Silvia Sedda  2 , Sanaa Choufani  3 , Isadora Asunis  1 , Giorgia Gemma  4   5 , Antonio Ammendola  5   6 , Arteen Torabi-Marashi  7 , Eleonora di Venere  5 , Gabriella Maria Squeo  5 , Vincenzo Rallo  1 , Maria Giuseppina Marini  1 , Paolo Moi  1   2 , Salvatore Savasta  2 , Rosanna Weksberg  3 , Giuseppe Merla  4   5 , Andrea Angius  1
Affiliations
Case Reports

A Novel Intronic Variant in the KH3 Domain of HNRNPK Leads to a Mild Form of Au-Kline Syndrome

Maura Mingoia et al. Clin Genet. 2025 Nov.

Abstract

Despite the massive adoption of sequencing technologies, disease-specific diagnosis remains challenging, particularly for genes with highly homologous pseudogenes like HNRNPK. Pathogenic HNRNPK variants cause Au-Kline syndrome (AKS), a neurodevelopmental disorder with malformations and distinctive facial features. We validated a novel de novo HNRNPK intronic variant (c.1192-3 C>A, p.Leu398ValfsTer21) in a patient previously misdiagnosed with Kabuki Syndrome (KS). By combining sequencing, in vitro splicing assays, molecular modelling, and protein function analysis, we characterised the molecular defect. A unique DNA methylation (DNAm) signature was recently identified in AKS, with missense variants showing an intermediate DNAm pattern, suggesting an epi-genotype-phenotype correlation linked to milder clinical features. The DNAm signature is a valuable tool for variant interpretation, especially in unclear AKS cases. We demonstrate that two independent approaches-functional characterisation and DNAm evaluation-confirmed a partial loss of HNRNPK function and validated an AKS diagnosis with a mild phenotype. Our findings highlight that a multidisciplinary approach integrating genomic and epigenomic analyses with functional studies and clinical assessment significantly improves rare disease diagnosis.

Keywords: Au‐Kline; DNA methylation; HNRNPK; alternative splicing; loss of function.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

FIGURE 1
FIGURE 1
Clinical and genetical features. (A) Patient facial appearance. (B) Family tree and DNA electropherograms. Red arrow indicates C > A de novo variant. (C) Schematic representation of the HNRNPK gene. Variant positions based on NM_002140.4. Arrows indicate DNAm signature evaluation for AKS variants: Black, positive signature; Blue, intermediate signature; Grey, positive/intermediate signature; Green, clinically confirmed AKS, no signature; Red, our de novo intermediate signature; According to 2 and 11 references.
FIGURE 2
FIGURE 2
Analysis of alternative splicing. (A) Schematic representation of the synthesis of the HNRNPK minigene into the pcDNA3.1 vector. (B) Electrophoresis of the cDNA splicing event of the wt (lane C) and mutated (lane A) minigene. (C) Sanger sequencing of the HNRNPK minigene. Upper, splicing of minigene c.1192‐3 A; middle, canonical wt splicing; lower, mutated alternative splicing. (D) RTqPCR of the HNRNPK.
FIGURE 3
FIGURE 3
Modelling and functional analysis. (A) Three‐dimensional structure of wild‐type KH3 domain (387‐451aa) and (B) mutated (387‐397 aa + 21 mutated aa). (C) Western blot of wt and mutated HNRNPK protein in HeLa cells. (D) Luciferase 3'UTR expression of pmiR‐GLO‐ALOX15 in HeLa cells.
See this image and copyright information in PMC

References

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