NFIX missense variants that disrupt the β-hairpin loop result in a severe form of Malan syndrome in adolescence with rapidly evolving scoliosis and muscle wasting
In vitro functional studies support variant-specific disruption of DNA binding, providing a mechanistic basis of genotype-phenotype correlations and informing prognosis, clinical surveillance, and therapy development.
Abstract
Purpose: Pathogenic variants in NFIX cause Marshall-Smith syndrome and Malan syndrome (MALNS). We identified a severe subtype of MALNS characterized by adolescent-onset musculoskeletal deterioration and investigated functional consequences of underlying variants. Methods: Clinical data were collected from seven individuals with pathogenic NFIX variants. Wild-type and mutated recombinant NFIX DNA-binding domains (DBDs) were evaluated using biochemical, structural, and DNA-binding assays. Results: Six individuals carrying R116W, R116P, K125E, or G147E NFIX substitutions developed progressive muscle wasting, markedly reduced body mass index, and rapidly progressive scoliosis after the typical childhood features of MALNS; two died from disease-related complications. A seventh individual with R116G did not develop this severe phenotype. Functional studies on recombinant NFIX DBDs showed complete or near-complete loss of DNA-binding activity for R116W, R116P, K125E, and G147E despite preserved protein folding, consistent with disrupted DNA recognition and a potential dominant-negative mechanism. In contrast, R116G exhibited a 7.7{degrees}C decrease in thermal stability, which may support haploinsufficiency mediated by protein degradation. Conclusion: Specific NFIX missense variants define a severe subtype of MALNS associated with progressive musculoskeletal deterioration. In vitro functional studies support variant-specific disruption of DNA binding, providing a mechanistic basis of genotype-phenotype correlations and informing prognosis, clinical surveillance, and therapy development.
Purpose: Neurofibromatosis type 2-related schwannomatosis (NF2-SWN) is an autosomal dominant tumor syndrome with complete penetrance and variable expressivity. Underlying patterns of disease burden and severity are largely unexplained. Methods: We comprehensively phenotyped 168 NF2-SWN patients over a mean duration of 4.5 years. We used a custom sequencing panel of NF2 and schwannomatosis genes to identify germline (n=166) and somatic variants in tumors (n=37). An optimized composite severity (CSS) score based on clinical and radiological data was created to analyze the effect of genetic variants on phenotype. Results: We found significant variable expressivity not explainable by demographic variables. Germline variants included premature termination (42%), splice-site (18%), and large deletions (16%). The CSS successfully predicted worsening clinical function in patients. Unsupervised clustering of clinical data revealed distinct phenotypic clusters that corresponded to CSS. Mosaicism, however, was not associated with CSS or any other disease severity marker. CSS was significantly associated with germline variant location along the NF2 locus. Specifically, FERM-F1 and the -helical variants were associated with increased disease severity. Within tumors, germline variants with severe effects on merlin acquired milder somatic second-hits at the NF2 locus. Conclusion: We identified a second-hit modifier to the Mendelian first-hit: severe germline variants were associated with milder somatic variants, and vice versa. This phenomenon partly explains the variable expressivity in NF2-SWN.
N. Ravindra, David T. Asuzu, Emma C. Celano et al.· medRxiv· 0 citations
The first functional characterization of the cardiomyopathy-associated SMYD1 N101S variant identified in a child with severe infantile cardiomyopathy is provided, establishing a mechanistic link between SMYD1 dysfunction and infantile cardiomyopathy and highlighting the importance of integrating genomic and functional approaches in rare cardiovascular disease.
Marta W. Szulik, Clint Gwynn, Magnus Creed et al.· bioRxiv· 0 citations
In this study, pathogenic FBN1 variants were identified in three patients, thereby confirming the clinical diagnosis of MFS and contributing two novel variants to the FBN1 variant repository, and provide a comparative assessment of genotype-phenotype correlations.
Xing Zhao, L. Yuan, Yan Sun et al.· Clinica chimica acta; intern...· 0 citations
Pyle disease is a rare autosomal recessive bone dysplasia characterized by a modeling defect of the tubular bones resulting in metaphyseal widening and cortical thinning, and bone fragility with fractures. Bi-allelic truncating variants in SFRP4 have been described as the primary molecular cause for Pyle disease. One report described a family with Pyle disease with compound heterozygous missense variants in SFRP4, yet no functional studies were performed. SFRP4 encodes the secreted Frizzled related protein 4 (sFRP4), which acts as a negative regulator of WNT signaling through the binding with WNT ligands or Frizzled receptors. We describe an adult woman with clinical and radiographic features of Pyle disease including wide metaphyses (Erlenmeyer-flask deformity), cortical thinning and multiple fractures. Genetic testing led to the identification of a novel homozygous missense variant (c.314G>C, p.(Arg105Pro)) in SFRP4, which is localized in the WNT-binding domain of sFRP4, similar to the previously reported missense variants (p.(Ala54Asp); p.(Cys125Ser)). Luciferase reporter experiments demonstrated that the p.(Arg105Pro) variant impairs the inhibitory potential of sFRP4 on WNT/β-catenin signaling. Similar impairment was also observed for the previously reported p.(Ala54Asp) and p.(Cys125Ser) variants. We therefore confirm that missense variants in SFRP4 may have a similar detrimental effect on sFRP4 function as truncating variants. This expands the mutational spectrum of Pyle disease and may improve the diagnosis of future families with this rare disorder. It also highlights the critical role of the WNT-binding domain of sFRP4 to act as a WNT signaling inhibitor.
Lex Magnus, E. Hordyjewska-Kowalczyk, Anna Sowińska-Seidler et al.· Bone· 0 citations
Abstract Robinow Syndrome is a genetically heterogeneous, rare skeletal disorder characterized by craniofacial and limb defects. All 7 causative genes lie in the Wingless-related Integration site-1 (WNT) pathway. Here we study the pathogenesis of DVL1 (Dishevelled 1), the most commonly affected gene, where variants cause a frameshift that replaces the C terminus with a novel peptide. We compared phenotypes caused by DVL11519ΔT to the effects of wtDVL1 or DVL1 with a stop codon at position 1519. Misexpression of DVL11519ΔT in chicken embryos with an avian retrovirus, leads to increased width of the frontonasal mass similar to the facial phenotype in RS. Ultimately skeletogenesis is inhibited, which was verified in primary cultures of frontonasal mass mesenchyme. In luciferase assays carried out in facial mesenchyme, wtDVL1 activated canonical and JNK-PCP WNT signalling whereas the DVL11519* and the DVL11519ΔT variant had significantly lower signaling activity. These data confirm that the C-terminus plays an important role in WNT signal transduction and skeletogenesis. We also determined that there is mislocalization of the protein expressed from DVL11519ΔT in the nucleus while the other two constructs were expressed in the cytoplasm. Nuclear expression of DVL1 may alter transcription in RS. In complementary Drosophila experiments using a variety of readouts, only the DVL11519ΔT variant and not the 1519* impacted morphogenesis and signaling. This is the first study to show that the novel C-terminus of DVL1 is sufficient to interfere with the function of DVL1 protein expressed from the normal allele in heterozygous, autosomal dominant RS.
Shruti S. Tophkhane, G. Akarsu, S. Gignac et al.· Human Molecular Genetics· 0 citations
Combined structural and functional evidence indicates that this variant disrupts protein stability, activates ER stress, and impairs ER–mitochondria communication, supporting its potential pathogenic role.
Silvia Borecka, A. Zahradníková, Lukáš Varga et al.· Molecular Medicine· 0 citations