Fam20C is an atypical
kinase implicated in bio-mineralization and
phosphate homeostasis disorders, and has recently been shown to account for the activity of an orphan
enzyme ("genuine
casein kinase", G-CK) previously characterized for its ability to phosphorylate
casein and a plethora of secreted
proteins at
serine residues specified by the S-x-E/pS motif. Fam20C/G-CK activity is only appreciable in the presence of high Mn2+ concentration (>1 mM), and is negligible if Mn2+ is replaced by physiological Mg2+ concentrations. Here we show that
sphingosine (but not its
biological precursor
ceramide) not only stimulates several-fold Fam20C activity in the presence of Mn2+, but also confers a comparable activity to Fam20C assayed with Mg2+. Activation by
sphingosine is evident using a variety of substrates, and is accounted for by both higher Vmax and decreased KmATP, as judged from kinetics run with the β(28-40) substrate
peptide and a physiological substrate,
BMP-15.
Sphingosine also protects Fam20C from thermal inactivation. Consistent with the in vitro results, by treating Fam20C expressing HEK293T cells with
myriocin, a potent inhibitor of the
sphingosine biosynthetic pathway, the activity of Fam20C released into the
conditioned medium is substantially decreased corroborating the concept that
sphingosine (or related metabolite(s)) is a co-factor required by Fam20C to optimally display its
biological functions. None of the small molecule
kinase inhibitors tested so far were able to inhibit Fam20C. Interestingly however
fingolimod, an immunosuppressive
drug structurally related to
sphingosine, used for the treatment of
multiple sclerosis, is a powerful activator of Fam20C, both wild type and its pathogenic, loss of function, T268M mutant. This article is part of a Special Issue entitled: Inhibitors of
Protein Kinases.