Bioinformatic Evaluation of KLF13 Genetic Variant: Implications for Neurodevelopmental and Psychiatric Symptoms

Abstract

The Krüppel-like factor (KLF) family represents a group of transcription factors (TFs) performing different biological processes that are crucial for proper neuronal function, including neuronal development, synaptic plasticity, and neuronal survival. As reported, genetic variants within the KLF family have been associated with a wide spectrum of neurodevelopmental and psychiatric symptoms. In a patient exhibiting attention deficit hyperactivity disorder (ADHD) combined with both neurodevelopmental and psychiatric symptoms, whole-exome sequencing (WES) analysis revealed a de novo heterozygous variant within the Krüppel-like factor 13 (KLF13) gene, which belongs to the KLF family and regulates axonal growth, development, and regeneration in mice. Moreover, in silico analyses pertaining to the likely pathogenic significance of the variant and the impact of the mutation on the KLF13 protein structure suggested a potential deleterious effect. In fact, the variant was localized in correspondence to the starting residue of the N-terminal domain of KLF13, essential for protein-protein interactions, DNA binding, and transcriptional activation or repression. This study aims to highlight the potential involvement of the KLF13 gene in neurodevelopmental and psychiatric disorders. Nevertheless, we cannot rule out that excluded variants, those undetectable by WES, or the polygenic risk may have contributed to the patient's phenotype given ADHD's high polygenic risk. However, further functional studies are required to validate its potential contribution to these disorders.

Keywords: Krüppel-like factor; autosomal dominant inheritance; axonal growth; missense mutation; next-generation sequencing; transcription factors.

Figure 1

Detection of c.20T>G within KLF13 gene. (a) Depiction of the nucleotide sequence corresponding to the region where the mutation was identified within the KLF13 gene. Furthermore, the chromosomal localization of this gene is illustrated. Figure was modified from the UCSC genome database. The asterisk indicates the precise variant site. (b) Whole-exome sequencing (WES) results are presented using the Integrative Genomics Viewer (IGV) visualization tool. As shown in the picture, WES was carried out for the examined patient and both healthy parents. (c) Conventional Sanger sequencing was performed to highlight the c.20T>G variant identified by WES. In the electropherograms, the black, blue, green, and red profiles indicate nucleotides G, C, A, and T.

Figure 2

Structure prediction analysis and functional domains related to KLF13 protein. (a) Protein structure prediction related to the wild-type KLF13. Each functional domain is marked by different colors. (b) Focus on the wild-type valine residue at position 7, which did not engage in hydrogen bonds with other amino acids. (c) Mutated KLF13 protein. As predicted, the different structural protein folding as result of the mutation is evident. (d) Close-up of the mutated residue as a result of the missense mutation p.Val7Gly. (e) Domain organization patterns related to the KLF13 protein. The specific mutation site is indicated by the black arrow. The light blue asterisk in (b,d,e) indicates the precise position of the missense mutation. (a–d) were generated by UCSF ChimeraX software, while (e) was modified from Uniprot database.

Figure 3

Structure prediction analysis of the KLF13 protein, focusing on the amino acid residues from positions 3 to 19, revealed significant structural variation from the primary structure to an α helix. Notably, the N-terminal domain of Krüppel-like factor 13 (from residues 7 to 168) begins at the specific mutation site at amino acid 7. The colors used are consistent with the domain organization patterns shown in Figure 2. (a) Wild-type KLF13 protein. (b) Close-up of the wild-type protein segment from alanine 3 to serine 19. (c) Mutated KLF13 protein, with visibly different predicted protein folding compared to the wild type. (d) Close-up of the segment from alanine 3 to serine 19, highlighting the mutated glycine 7 (marked with an asterisk). The missense mutation is predicted to result in the formation of an α helix containing 17 hydrogen bonds. (a–d) were generated by UCSF ChimeraX software and subsequently modified.

Figure 4

Line plots generated with PONDR tool with VLXT score to assess the impact of the mutation on protein stability and flexibility. (a) The VLXT score (blue line) from PONDR analysis for the wild-type KLF13 protein indicates a high rate of structural order at the specific site (green line indicating valine at position 7), with scores lower than 0.5 (orange line). (b) The VLXT score from PONDR analysis for the mutated KLF13 protein shows a higher rate of disorder as a result of the mutation. This is evidenced by the values of the residues before the mutation site being higher than 0.5 (orange line).