posted on 2019-12-10, 14:59authored byChen Shen, Bin Li, Luisana Astudillo, Murray P. Deutscher, Melanie H. Cobb, Anthony J. Capobianco, Ethan Lee, David J. Robbins
The serine/threonine protein kinase casein kinase 1α
(CK1α)
functions as a negative regulator of Wnt signaling, phosphorylating
β-catenin at serine 45 (P–S45) to initiate its eventual
ubiquitin-mediated degradation. We previously showed that the repurposed,
FDA-approved anthelminthic drug pyrvinium potently inhibits Wnt signaling in vitro and in vivo. Moreover, we proposed
that pyrvinium’s Wnt inhibitory activity was the result of
its function as an activator of CK1α. An understanding of the
mechanism by which pyrvinium activates CK1α is important because
pyrvinium was given an orphan drug designation by the FDA to treat
familial adenomatous polyposis, a precancerous condition driven by
constitutive Wnt signaling. In the current study, we show that pyrvinium
stimulates the phosphorylation of S45 β-catenin, a known CK1α
substrate, in a cell-based assay, and does so in a dose- and time-dependent
manner. Alternative splicing of CK1α results in four forms of
the protein with distinct biological properties. We evaluated these
splice products and identified the CK1α splice variant, CK1αS,
as the form that exhibits the most robust response to pyrvinium in
cells. Kinetic studies indicate that pyrvinium also stimulates the
kinase activity of purified, recombinant CK1αS in vitro, increasing its catalytic efficiency (kcat/Km) toward substrates. These studies
provide strong and clear mechanistic evidence that pyrvinium enhances
CK1α kinase activity.