The CDKL5 kinase undergoes liquid–liquid phase separation driven by a serine-rich C-terminal region

This article uncovers the ability of the CDKL5 kinase to form intracellular condensates through liquid–liquid phase separation primarily mediated by a serine-rich low-complexity region. The loss of this region caused by neurodevelopmental disease-related mutations can alter CDKL5 localization and function. The CDKL5 gene encodes a protein kinase involved in nervous system development and function. Pathogenic variants in this gene can cause a severe neurodevelopmental CDKL5 deficiency disorder (CDD). The CDKL5 protein contains a catalytic N-terminal domain (NTD) and a less characterized C-terminal domain (CTD). We discovered that the CTD is a serine-rich low-complexity region driving liquid–liquid phase separation (LLPS), a biophysical process controlling protein localization and function, by which CDKL5 forms intracellular membraneless condensates. A CTD internal fragment (CTIF) plays a pivotal LLPS-promoting role, along with the distal portion of the protein. In CDD, transcripts carrying pathogenic nonsense or frameshift mutations introducing distal premature termination codons may escape nonsense-mediated decay, producing CDKL5 proteins with a variably truncated CTD. We found that two distal truncations, removing part of the CTIF and the downstream protein tail, significantly reduce CDKL5 LLPS and catalytic function. These findings demonstrate that CDKL5 undergoes LLPS, driven by a CTD region whose loss in distally truncated forms of the protein—by impairing LLPS and functional activity—may play a role in the molecular pathogenesis of CDD.