Assessment of paralogue annotation for improving diagnostic accuracy in CALM1, CALM2, and CALM3 genes

Figures

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  FIGURE 1. Methodological approach for paralogue annotation (PA) of CALM1, CALM2, and CALM3 variants. ClinVar and Mastermind databases were used to identify CALM1, CALM2, and CALM3 missense variants. Non-missense, duplicate, benign, and likely benign variants were removed. The two datasets were merged and manually curated according to ACMG guidelines based on evidence available for each individual gene prior to PA. The final dataset comprised 126 unique cDNA or 110 protein variants.

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  FIGURE 2. Number of pathogenic and VUS variants in CALM1, CALM2, and CALM3 genes in ClinVar (A) and Mastermind (B). Abbreviations: VUS, variants of uncertain significance; P, pathogenic variants reported as either pathogenic or likely pathogenic. (C) Conflicting classification of pathogenicity. Application of paralogue annotation (PA) to CALM1, CALM2, and CALM3 genes. Before and after PA, the use of PA led to a 111% increase in P across CALM1, CALM2, and CALM3 genes.

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  FIGURE 3. (A) 3D representation of C-lobe of the calmodulin (CaM) protein, highlighting the EF-hand 4 motif (labeled). Arrows point to the specific P variants reported in our dataset located at the specific locations colored in red. Key residues for protein function reported as P are shown with white side chains; dashed white lines indicate the distance (in angstroms, Å) between these residues and the bound calcium ion (Ca2+), potentially influencing CaM’s interaction with calcium and its downstream effects. (B) EF-hand 3 motif and variants located in this region. (C) Structure of the calmodulin protein in the holo form, a calcium-bound form essential for its function. Cyan spheres represent calcium ions (Ca2+). N-lobe (blue) and C-lobe (orange) are the functional domains of calmodulin, connected by the central tether (green). Notably, red spheres highlight the locations of pathogenic variants reported within the protein structure.