FGF12 Human

What is FGF12 and what is its role in human physiology?

FGF12 is a member of the fibroblast growth factor (FGF) homologous factor family, which includes FGF11, FGF12, FGF13, and FGF14. Unlike classical FGFs, FGF12 functions as an intracellular protein rather than a secreted growth factor. It interacts with the C-terminal domain of the alpha subunit of voltage-gated sodium channels (NaVs) 1.2, 1.5, and 1.6 .

FGF12's primary physiological role is to modulate neuronal excitability by delaying fast inactivation of these voltage-gated sodium channels, thereby promoting excitability . This function is critical for normal neuronal signaling and cardiac function. In the brain, FGF12 expression affects both excitatory and inhibitory neurons, maintaining the balance necessary for proper neurological function . In cardiac tissue, FGF12 appears to interact with proteins involved in energy metabolism and plays a role in regulating cardiomyocyte hypertrophy .

How are FGF12 variants associated with human diseases?

FGF12 variants are associated with several neurological disorders, primarily developmental and epileptic encephalopathy (DEE). Specific genetic alterations in FGF12 have different pathophysiological consequences:

The clinical presentations include tonic seizures, intellectual disability, speech problems, autistic features, and ataxia . Additionally, reduced FGF12 expression has been observed in patients with hypertrophic cardiomyopathy (HCM) .

What genomic approaches are most effective for detecting FGF12 variants?

The optimal detection of FGF12 variants requires a comprehensive approach due to the diverse nature of potential mutations:

For comprehensive detection of all possible FGF12 variants, researchers should consider using a combination of these techniques, particularly when investigating patients with neurological disorders and negative standard genetic testing results.

How can FGF12 expression be accurately measured in patient samples?

Measuring FGF12 expression presents several challenges due to its typically low expression levels in accessible tissues. Based on research methodologies, the following approaches are recommended:

• Droplet Digital PCR (ddPCR): Highly sensitive technique capable of detecting low expression levels of FGF12 in lymphoblastoid cell lines (LCLs). This method overcomes the limitations of regular real-time PCR for detecting the low levels of FGF12 expression in LCLs .

• Allele-Specific Expression (ASE) Analysis using ddPCR: Important for distinguishing expression from normal and variant alleles. This can be performed using TaqMan probes designed to distinguish between specific alleles .

• Cycloheximide (CHX) Treatment for NMD Assessment: Treatment of cell lines with CHX followed by RT-PCR and Sanger sequencing can help determine whether abnormal transcripts are subject to nonsense-mediated decay (NMD) .

• Complementary DNA (cDNA) Analysis: PCR of cDNA can detect both normal and aberrant transcripts, such as those containing duplicated exons .

• CUT&Tag Sequencing: Useful for investigating FGF12 binding to gene promoter regions, particularly relevant in studies of its role in cardiomyocyte hypertrophy .

When designing expression studies, researchers should be aware that FGF12 expression patterns differ significantly between tissue types, with neuronal tissues typically showing higher expression than lymphoblastoid cells used in most accessible patient samples.

What are the key molecular interactions of FGF12 in neurons and cardiomyocytes?

FGF12 exhibits tissue-specific interactions that contribute to its distinct functional roles:

In Neurons:

• Interacts with the C-terminal domain of voltage-gated sodium channels (NaVs) 1.2 and 1.6

• Modulates channel inactivation gating, affecting neuronal excitability

• Loss-of-function of FGF12 appears to impair the balance between excitatory and inhibitory neuronal activities, potentially leading to epilepsy

In Cardiomyocytes:

• Associates with proteins involved in energy metabolism

• Interacts with GATA binding protein 4 (GATA4) and mitogen-activated protein kinase 1/3 (MAPK1/3) in the perinuclear space

• Upon translocation to the nucleus in hypertrophic states, binds to the GATA4 promoter region, increasing its expression

• Interaction with NaV1.5 in cardiomyocytes impacts cardiac excitability

• Inhibits GATA4 and MAPK1/3 phosphorylation under normal conditions, but this inhibition is lost upon nuclear translocation in hypertrophy

Understanding these interactions provides crucial insights into the pathophysiological mechanisms of both neurological disorders and cardiomyopathies associated with FGF12 dysfunction.

How does cellular localization affect FGF12 function?

FGF12's function is significantly influenced by its subcellular localization:

• Perinuclear Localization (Normal Physiological State):

  • Under normal conditions, FGF12 predominantly localizes to the perinuclear space

  • In this location, it interacts with GATA4 and MAPK1/3, inhibiting their phosphorylation

  • This inhibition appears to prevent hypertrophic responses in cardiomyocytes

• Nuclear Translocation (Pathological State):

  • In hypertrophic cardiomyocytes, FGF12 shifts from the perinuclear space into the nucleus

  • Upon nuclear translocation, FGF12 binds to the GATA4 promoter region

  • This binding increases GATA4 expression and activates the ERK1/2-pGATA4 pathway genes associated with hypertrophy

  • Deletion of the nuclear localization signal (NLS) in FGF12 results in decreased GATA4 phosphorylation, suggesting that nuclear translocation is essential for FGF12's role in promoting hypertrophy

These findings suggest that the subcellular trafficking of FGF12 represents a critical regulatory mechanism that determines its functional impact on cellular processes, particularly in cardiomyocyte hypertrophy.

What model systems are most appropriate for studying FGF12 function?

Multiple model systems have been effectively utilized to study FGF12 function, each with specific advantages for addressing different research questions:

Cellular Models:

• Lymphoblastoid Cell Lines (LCLs): Useful for gene expression studies in patient samples, though with limitations due to low FGF12 expression levels

• Human induced Pluripotent Stem Cell-derived Cardiomyocytes (hiPSC-CMs): Valuable for studying FGF12's role in cardiac function and hypertrophy

• Heterologous Expression Systems: Important for electrophysiological studies of FGF12 interactions with sodium channels

Animal Models:

• Drosophila: Effective for in vivo functional analysis of FGF12 variants, particularly for assessing loss-of-function effects

• Transgenic Mouse Models: Including MYH7 R403Q and MYBPC3 c.790G>A mouse models for hypertrophic cardiomyopathy

• Transverse Aortic Constriction (TAC) Mouse Model: Useful for studying cardiac hypertrophy mechanisms related to FGF12

Human Samples:

• Patient-derived Tissues: Provide direct evidence of FGF12's role in human pathophysiology

• Post-mortem Brain Tissue: Important for studying FGF12 expression and localization in the context of neurological disorders

When selecting a model system, researchers should consider the specific aspect of FGF12 biology they aim to investigate and the relevance of that model to human pathophysiology.

What methodological approaches are recommended for investigating FGF12's role in epilepsy?

To comprehensively investigate FGF12's role in epilepsy, a multi-faceted approach is recommended:

• Genetic Screening:

  • Long-read whole genome sequencing to detect structural variations

  • Exome sequencing for identification of single-nucleotide variants

  • Segregation analysis in families to confirm biallelic inheritance patterns

• Expression Analysis:

  • Droplet digital PCR for sensitive quantification of FGF12 expression levels

  • Allele-specific expression analysis to distinguish expression from normal and variant alleles

  • Analysis of nonsense-mediated decay through cycloheximide treatment

• Functional Studies:

  • Electrophysiological recordings of sodium current in the presence/absence of FGF12 proteins

  • Assessment of neuronal excitability in models expressing wild-type or variant FGF12

  • Drosophila in vivo functional analysis for studying the effects of specific variants

• Structural Analysis:

  • In silico structural modeling to predict the impact of variants on protein function

  • Analysis of evolutionary conservation of affected amino acid residues

• Clinical Correlation:

  • Detailed phenotyping of patients with FGF12 variants

  • Brain imaging (MRI) to assess structural abnormalities such as cerebral or cerebellar atrophy

  • Electroencephalography (EEG) to characterize seizure patterns

This integrated approach enables researchers to establish causality between FGF12 variants and epilepsy phenotypes while elucidating the underlying pathophysiological mechanisms.

What are potential therapeutic targets related to FGF12 dysfunction?

Based on current understanding of FGF12 pathophysiology, several potential therapeutic targets emerge:

• For Gain-of-Function FGF12-Related Epilepsy:

  • Direct targeting of the FGF12-sodium channel interaction to normalize channel inactivation

  • Modulation of sodium channel function to counteract the enhanced excitability caused by FGF12 variants

  • Development of antisense oligonucleotides to reduce expression of gain-of-function FGF12 variants

• For Loss-of-Function FGF12-Related Epilepsy:

  • Approaches to enhance residual FGF12 function or expression

  • Targeting compensatory mechanisms to restore balance between excitatory and inhibitory neuronal activities

  • Gene therapy approaches to introduce functional FGF12 in affected tissues

• For FGF12-Related Cardiomyopathy:

  • Inhibition of FGF12 nuclear translocation to prevent binding to the GATA4 promoter

  • Modulation of the ERK1/2-pGATA4 pathway to counteract hypertrophic responses

  • Therapeutic strategies targeting the interaction between FGF12 and GATA4 or MAPK1/3

These therapeutic approaches require further validation in appropriate model systems before translation to clinical applications.

How might contradictions in FGF12 research findings be resolved?

Several apparent contradictions exist in the current literature on FGF12, requiring careful consideration and potential resolution strategies:

• Contradictory Effects of FGF12 Variants:

  • Both gain-of-function and loss-of-function variants in FGF12 appear to cause epilepsy

  • Resolution approach: Investigate tissue-specific effects and potential differences in the balance of excitatory versus inhibitory neuronal activities affected by different variants

• Inconsistent Phenotypic Effects of Similar Variants:

  • Heterozygous partial duplication of FGF12 has been reported in DEE individuals, but the carrier mother with a similar duplication in the current study was unaffected

  • Resolution approach: Examine differences in duplication sizes and exons involved, or investigate potential hidden structural variations in the "normal allele"

• Limitations of Model Systems:

  • Gene expression analysis using lymphoblastoid cell lines may not fully represent FGF12 function in neurons or cardiomyocytes

  • Resolution approach: Compare findings across multiple model systems, including induced pluripotent stem cell-derived neurons and cardiomyocytes that more closely represent the relevant tissues

• Methodological Discrepancies:

  • Different detection methods (LRWGS vs. OGM) yield varying results regarding the sizes and breakpoints of structural variations

  • Resolution approach: Use multiple complementary technologies and validate findings with orthogonal methods

Addressing these contradictions requires careful experimental design, replication in multiple model systems, and consideration of tissue-specific and developmental context-dependent effects of FGF12.