Function of PHEX mutations p.Glu145* and p.Trp749Arg in families with X-linked hypophosphatemic rickets by the negative regulation mechanism on FGF23 promoter transcription

Cell Death Dis. 2022 Jun 2;13(6):518. doi: 10.1038/s41419-022-04969-5.

PMID: 35654784

Yu-Mian Gan, Yan-Ping Zhang, Dan-Dan Ruan

Highlights: This study verified the pathogenicity of the two variants and revealed the possible regulatory mechanism between PHEX and FGF23.

Abstract

Background: PHEX (NM 000444.5) mutations cause the circulating fibroblast growth factor 23 (FGF23) levels to increase in individuals with X-linked hypophosphatemic rickets (XLH). Rickets and reduced bone mineralization come from impaired renal tubular phosphate resorption.

Results: Two PHEX pathogenic mutations were identified by phenotypic-genetic linkage analysis in two XLH families: c.433 G > T, p.Glu145* in exon 4 and c.2245 T > C, p.Trp749Arg in exon 22. p.Glu145* and p.Trp749Arg mutants and secretory PHEX (secPHEX) have altered location and decreased expression, according to immunofluorescence. The wild-type PHEX, secPHEX, and FGF23 proteins were dispersed in the cell membrane or endoplasmic reticulum in a co-transfected HEK293T cell model, whereas the mutant was found in the nuclear membrane and cytoplasm. p.Glu145* qPCR results showed lower PHEX and secPHEX mRNA expression in cells, but no difference in p.Trp749Arg mRNA expression. The activity of the intracellular PHEX endopeptidase was reduced by both mutations. In single-transfected PHEX and secPHEX cells, Western blot analysis revealed decreased mutant and secPHEX protein expression and no FGF23 protein expression. PHEX and secPHEX mutation increased FGF23 production in cells that also received FGF23 co-transfection. PHEX's impact on the FGF23 promoter was discovered using a dual-luciferase reporter gene. The mutant firefly luciferase had considerably higher activity than the wild type after PHEX overexpression, according to the dual-luciferase reporter gene.

Conclusion: The regulatory mechanism between PHEX and FGF23 is still unclear, but it is found that PHEX is a direct transcriptional inhibitor of FGF23 and affects the expression of FGF23. The two variants' pathogenicity was confirmed by this investigation, which also identified a potential regulatory link between PHEX and FGF23.

Keywords: PHEX, X-linked hypophosphatemic rickets, XLH