HRI Kinase is Involved in the Development of Dental Fluorosis

Dental fluorosis affects approximately one quarter of the American population but the mechanism behind it remains poorly understood.  We have previously shown the phosphorylation of the eIF2α ribosomal subunit in ameloblasts of fluoride-treated wild-type (WT) mice.  Phosphorylation of eIF2α decreases overall protein synthesis via abrogated binding of the initiator methionine tRNA to the ribosome.  Here we show the involvement of the eIF2α-kinase heme-regulated inhibitor (HRI) during the response to fluoride in vitro and propose that HRI inhibits protein removal from developing enamel by phosphorylation of eIF2α. 

Objective: To identify the upstream kinase(s) involved in fluoride–mediated eIF2α phosphorylation and to characterize the stress response pathway involved in the development of dental fluorosis.

Method: In vitro cellular proliferation assays of kinase-null murine embryonic fibroblast (MEF) cell lines and protein secretion assays of RNA-interfered ameloblast-derived LS8 cells were performed in both the presence and absence of sodium fluoride.

Result: We show the presence of HRI mRNA in ameloblast-lineage cells (ALC), LS8 and MEF cell lines as well as in both secretory- and maturation-stage enamel organs from WT mice.  Cellular proliferation of MEF cells was affected in a dose-dependent manner by fluoride in a similar manner in both eIF2α-phosphorylation negative and HRI-null backgrounds; this suggests HRI and eIF2α operate in the same pathway in response to fluoride.  Luciferase secretion was decreased in LS8 cells in the presence of fluoride in control samples but was affected to a lesser extent in the HRI-knockdown cells implying HRI is responsible for the eIF2α-mediated protein synthesis decrease in response to fluoride.

Conclusion: Our data suggest HRI is the kinase responsible for eIF2α phosphorylation during fluoride exposure in vivo and suggests that retention of protein in fluorotic enamel is caused by the decreased synthesis of proteins, such as the enamel matrix protease kallikrein-4.  Supported by NIDCR grant DE018106.