Human FGF6 mRNA

Standard analyses applied to genome-wide association data are well designed and capable of detecting additive effects of moderate magnitude. However, standard genome-wide association study (GWAS) analyses are often poorly powered to identify recessive diploid effects. Here, researchers propose and perform a gene-based test for compound heterozygosity to reveal additional genes underlying complex diseases. Applying this approach to iron overload, a strong association signal was found between the fibroblast growth factor encoding gene FGF6 and hemochromatosis in a central Wisconsin population. Functional validation demonstrated that the fibroblast growth factor 6 protein (FGF-6) regulates iron homeostasis and induces transcriptional regulation of hepcidin. Furthermore, specific FGF6 variants identified differentially affect iron metabolism. Furthermore, FGF6 downregulation is associated with dysfunctional iron metabolism in systemic sclerosis and cancer cells. Using a recessive diploid approach revealed a new hemochromatosis susceptibility gene and expanded the understanding of the mechanisms of iron metabolism.

To investigate the potential mechanisms of the association between FGF-6 and iron metabolism, the researchers evaluated the effects of FGF6 on iron uptake and expression of iron metabolism genes in HepG2, HCT8, HCT116, 786-O, and HFF1 cells. By culturing cells and detecting iron using ferrozine assays, the researchers found that total intracellular iron concentrations were significantly reduced in HepG2, 786-O, HCT8, HCT116, and HFF-1 cells when treated with active FGF-6 protein in a dose-dependent manner. To test the effects of FGF6 on the expression of a panel of genes involved in iron metabolism (HAMP, HDAC2, HMOX1, TFRC, and HEPH), HepG2 cells were subjected to treatment from control or FGF-6 protein and FGF6 mRNA or control and mRNA expression relative to GAPDH was measured in the 5 iron-metabolism genes. RT-PCR analysis showed that HAMP and HDAC2 mRNA levels were significantly increased in HepG2 cells after the introduction of FGF-6 active protein compared with the PBS control group (Figure 1A). FGF6 plasmid transfection significantly increased the levels of HAMP, HDAC2, and HMOX1 in HepG2 cells compared with the vector without FGF6 introduction, while the level of TFRC was significantly decreased (Figure 1B). The expression of HEPH did not change with the introduction of FGF6 plasmid or FGF-6 protein, suggesting that the effect of FGF-6 may be independent of HEPH (Figure 1A-B).

Figure 1. (A) The effect of FGF-6 active protein treatment on mRNA expression of several iron metabolism genes in HepG2 liver hepatocellular carcinoma cell culture media compared with control. (B) Iron-metabolism gene expression changes with FGF6 mRNA transfection in the HepG2 cell culture media after 24 hours. (Guo S, et al., 2019)