Browsing by Author "Pitt, Geoffrey S"

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The non-secreted fibroblast growth factor (FGF) homologous factor (FHF) FGF13 is a noncanonical FGF with identified roles in neuronal development, pain sensation, and cardiac physiology, but recent reports suggest broader roles. The in vivo functions of FGF13 have not been widely studied. In this study, we have generated a global heterozygous Fgf13 knockout mouse model. In these animals, we observed hyperactivity and accompanying reduced core body temperature in mice housed at 22 °C. In mice housed at 30 °C (thermoneutrality) we observed development of a pronounced obesity. Defects in thermogenesis and metabolism were found to be due to impaired central nervous system regulation of sympathetic activation of brown fat. Neuronal and hypothalamic specific ablation of Fgf13 recapitulated weight gain at 30 °C. In global heterozygous animals, norepinephrine turnover in brown fat was reduced at both housing temperatures, while direct activation of brown fat by a β3 agonist showed an intact response. Further, we found that FGF13 is a direct regulator of NaV1.7, a hypothalamic Na+ channel associated with regulation of body weight. Our data expand the physiologic roles for FGF13, and enhance the understanding of the multifunctional FHFs.

Fibroblast growth factor (FGF) homologous factors (FHFs, FGF11-14) are a family of FGFs that are not secreted, nor activate FGF receptors. Instead, they remain intracellular and bind to the voltage-gated Na+ channel C-terminus and modulate function. FGF14 is a locus for the neurodegenerative disease spinocerebellar ataxia 27 and the disease has been attributed to decreased neuronal excitability from changes in Na+ channel function. However, several lines of evidence, including data from heterologous expression systems and the distribution of FGF13 within the ventricular cardiomyocyte suggested that it also modulates the Ca_V1.2 voltage-gated Ca2+ channel. The central hypothesis to this study is that FHFs modulate both voltage-gated Na+ and Ca2+ channel channels in the ventricular cardiomyocyte and therefore are loci for cardiac arrhythmia. Using an adult ventricular cardiomyocyte system with adenoviral gene transfer, we manipulated the levels of FGF13 in the cell and performed electrophysiology, biochemistry and immunocytochemistry to analyze the effects on voltage-gated Ca2+ channel channel localization and function. We showed that FGF13 is in complex with Junctophilin-2 and modulates Ca_V1.2 current density and localization to the t-tubule, leading to changes in Ca2+ channel-induced Ca2+ channel release and ultimately a shortened ventricular action potential. Through collaboration with the Mayo Clinic, a mutation in FGF12, the most highly expressed FHF in human ventricle was found in a patient with Brugada syndrome. Using similar methodology, we determined that this mutation results specifically in a Na_V1.5 loss of function without affecting Ca_V1.2 function, resulting in a Brugada-like ventricular action potential. This data shows that FHFs are potent modulators of multiple ion channels and novel arrhythmogenic loci.

Fibroblast growth factor homologous factors (FHFs) are non-canonical members of the fibroblast growth factor family (FGF11-14) that were initially discovered to bind and regulate neuronal and cardiac voltage-gated Na+ channels. Loss-of-function mutations that disrupt interaction between FHFs and Na+ channels cause spinocerebellar ataxias and cardiac arrhythmias such as Brugada syndrome. Although recent studies in brain of FHF knockout mice suggested novel functions for FHFs beyond ion channel modulation, it is unclear whether FHFs in the heart serve additional roles beyond regulating cardiac excitability. In this study, we performed a proteomic screen to identify novel interacting proteins for FGF13 in mouse heart. Mass spectrometry analysis revealed an interaction between FGF13 and a complex of cavin proteins that regulate caveolae, membrane invaginations that organize protective signaling pathways and provide a reservoir to buffer membrane stress. FGF13 controls the relative distribution of cavin 1 between the plasma membrane and cytosol and thereby acts as a negative regulator of caveolae. In inducible, cardiac-specific Fgf13 knockout mice, cavin 1 redistributed to the plasma membrane and stabilized the caveolar structural protein caveolin 3, leading to an increased density of caveolae. In a transverse aortic constriction model of pressure overload, this increased caveolar abundance enhanced cardioprotective signaling through the caveolar-organized PI3 kinase pathway, preserving cardiac function and reducing fibrosis. Additionally, the increased caveolar reserve provided mechanoprotection, as indicated by reduced membrane rupture in response to hypo-osmotic stress. Thus, our results establish FGF13 as a novel regulator of caveolae-mediated mechanoprotection and adaptive hypertrophic signaling, and suggest that inhibition of FHFs in the adult heart may have cardioprotective benefits in the setting of maladaptive hypertrophy.

Fibroblast growth factor homologous factors (FHFs) are noncanonical members of the fibroblast growth factor family (FGFs, FGF11-FGF14) that bind directly to voltage gated sodium channels (VGSCs), thereby regulating channel activity and consequently neuronal excitability. Mutations in FGF14 cause spinocerebellar ataxia while FGF13 is a candidate for X-linked mental retardation. Since FGF13 and FGF14 are nearly identical within their respective VGSC-interacting domains, those distinct pathological consequences have generally been attributed to regional differences in expression. I have shown that FGF13 and FGF14 have non-overlapping subcellular distributions and biological roles even in hippocampal neurons where both are prominent. While both FHFs are abundant in the axon initial segment (AIS), only FGF13 is observed within the soma and dendrites. shRNA knockdown and rescue strategies showed that FGF14 regulates axonal VGSCs, while FGF13 only affects VGSCs in the somatodendritic compartment. Thus, FGF13 and FGF14 have nonredundant roles in hippocampal neurons, with FGF14 acting as an AIS-dominant positive regulator and FGF13 serving as a somatodendritic negative regulator. Both of these FHFs also perform important non-VGSC regulatory roles. FGF14 is a novel potassium channel regulator, which binds to KCNQ2 and regulates both localization and function. FGF14 is also capable of serving as a “bridge” between VGSCs and KCNQ2 thus implicating it as a broad organizer of the AIS. FGF13, on the other hand is involved in a new form of neuronal plasticity called axon initial segment structural plasticity. Knockdown of FGF13 impairs AIS structural plasticity and reduces L-type CaV current through channels known to be important to this new form of plasticity. Both of these novel non-VGSC roles are specific to the FHF in question because FGF13 does not regulate KCNQ2 whereas FGF14 knockdown does not affect AIS position. These data imply wider roles for FHFs in neuronal regulation that may contribute to differing roles in neural disease.