IONIC REGULATION OF NEK7 AND DDX3X PROTEINS

NLRP3 inflammasome is an intracellular protein complex that drives the activation of
inflammatory caspases. Ionic flux events of potassium, sodium, calcium, and chloride
ions are implicated in activating the NLRP3 inflammasome. Consequently, dysfunction
of its activation is associated with a variety of human diseases including neurological
disorders. We have investigated the effect of the chloride ion in the function of two
NLRP3 binding partners that drive the NLRP3 inflammasome activation: NEK7, a
serine/threonine kinase, and DDX3X, a DEAD-box RNA helicase. We verified the
presence of chloride ions in the RNA and ATP binding region of DDX3X in a previously
reported crystal structure of human DDX3X. Importantly, two intellectual disability (ID)-
linked DDX3X mutations, R326H and R475G, localize to this region. Using biophysical
assays, we demonstrated that both DDX3X and NEK7 interact with chloride ions. Our
biochemical experiments showed that DDX3X ATPase and NEK7 autophosphorylation
activities are independently impaired by the chloride ion. We are now investigating
how disease mutants of DDX3X respond to chloride. Together, our results will provide
new insights into the molecular mechanisms associated with chloride efflux and
NLRP3 inflammasome activation.