10779/rcsi.10790825.v1 Hans-Georg König Hans-Georg König Orla Watters Orla Watters Sinéad Kinsella Sinéad Kinsella Mohammed Ameen Mohammed Ameen Beau J Fenner Beau J Fenner Jochen H.M. Prehn Jochen H.M. Prehn A constitutively-active IKK-complex at the axon initial segment. Royal College of Surgeons in Ireland 2019 Animals Ankyrins Aspirin Axon Initial Segment Calbindins Cerebral Cortex Dose-Response Relationship Drug Embryo Mammalian Enzyme Inhibitors Flow Cytometry Green Fluorescent Proteins I-kappa B Kinase I-kappa B Proteins Ligation Mice Inbred C57BL Neurons Phosphorylation RNA Small Interfering Rats Serine Signal Transduction Time Factors Transfection Physiology Medical Physics 2019-11-22 16:56:46 Journal contribution https://repository.rcsi.com/articles/journal_contribution/A_constitutively-active_IKK-complex_at_the_axon_initial_segment_/10790825 <p>BACKGROUND: Previous studies provided evidence for an accumulation of IκB-kinase (IKK) α/β at the axon initial segment (AIS), a neuronal compartment defined by ankyrin-G expression. Here we explored whether the presence of the IKK-complex at the AIS was associated with the activation of IKK signaling at this site.</p> <p>METHODS AND RESULTS: Proximity-ligation assays (PLAs) using pan-IKKα/β, phospho-IKKα/β-specific as well as ankyrin-G specific antibodies validated their binding to proximal epitopes in the AIS, while antibodies to other phosphorylated signaling proteins showed no preference for the AIS. Small-hairpin mediated silencing of IKKβ significantly reduced anti-phospho-IKKα/β-immunoreactivities in the AIS. ank3 gene-deficient cerebellar Purkinje cells also exhibited no phosphorylated IKKα/β at the proximal region of their axons. Transient ankyrin-G overexpression in PC12 cells augmented NF-κB transactivation in an ankyrin-G death-domain dependent manner. Finally, small molecule inhibitors of IKK-activity, including Aspirin, inhibited the accumulation of activated IKK proteins in the AIS.</p> <p>CONCLUSION: Our data suggest the existence of a constitutively-active IKK signaling complex in the AIS.</p>