Pseudokinase-converting mutation in protein kinase C alpha drives chordoid glioma by pathway rewiring.

Abstract

Chordoid glioma (ChG) is a rare, low-grade brain tumor characterized by a novel recurrent point mutation, D463H, in the kinase domain of protein kinase C alpha (PKCα). The mutation is invariably an Asp to His substitution, suggesting a unique function beyond catalytic inactivation associated with other cancer-associated PKCα mutations. Here, we show that this mutation converts PKCα into a pseudokinase, abolishing catalytic activity, and, additionally, confers novel scaffolding functions. Activity assays in vitro and in cellulo revealed that PKCα_D463H is catalytically inactive and functions as a dominant-negative to suppress endogenous PKC activity. Molecular dynamics simulations predicted that mutation to His, but not Asn, not only destabilizes the active site, but stabilizes the substrate-binding helices in the kinase C-lobe to potentially promote aberrant interactions. Supporting this, phosphoproteomic, proximity labeling, and coimmunoprecipitation mass spectrometry data from cells overexpressing PKCα_D463H identified both altered phosphorylation of substrates and binding to multiple proteins involved in cell-cell junctions compared to WT enzyme. Last, single nuclei RNAseq established that ChG derives from specialized tanycytes. Our data reveal that this disease-defining, fully penetrant mutation converts PKCα into a pseudokinase with novel scaffold functions that uniquely rewire the cellular interactome to impair cell junction function.