Deciphering Silence: Functional Studies of GCK Synonymous and Nonsense Variants and Their Importance in Understanding...

Background: The most common form of monogenic diabetes is maturity onset diabetes of the young (MODY). This study investigates the molecular basis of MODY type 2 (GCK-MODY) in a group of Italian patients, focusing on the functional characterization of a synonymous variant, c.579G>T (p.Gly193Gly), in the glucokinase gene (GCK). Methods: Clinical evaluation and genetic analysis, including whole exome sequencing and Sanger sequencing, were used to identify the variant in GCK, then functional studies using a minigene approach allowed the functional characterization. Results: This study identified the synonymous variant, along with a nonsense mutation, c.859C>T (p.Gln287Ter), in GCK in two Italian patients. Minigene approach demonstrated that the synonymous variant disrupts splicing at the exon 5 boundary, leading to a frameshift and premature stop codon. Similarly, the nonsense mutation also altered splicing, exacerbating the molecular defect. Conclusions: These findings highlight the importance of functional assays, particularly minigene studies, in interpreting the pathogenicity of synonymous and nonsense variants, especially in genes like GCK where splicing alterations can significantly impact protein function. This study underscores the clinical utility of targeted genetic screening for personalized diabetes management.