Inositol hexakisphosphate kinase‐2 in cerebellar granule cells acts through protein 4.1N to regulate Purkinje cell morphology and motor coordination

Inositol hexakisphosphate kinases (IP6Ks) are responsible for the synthesis of the energy‐rich Inositol pyrophosphates (PP‐IPs) in mammals and regulate cellular functions including chemotaxis, telomere length, endocytic trafficking, exocytosis as well as apoptosis. Among such pyrophosphates, diphosphoinositol pentakisphosphate (IP7) and bis‐diphosphoinositol tetrakisphosphate (IP8) have been extensively characterized. IP7 is produced in mammals by a family of inositol hexakisphosphate kinases, namely IP6K1, IP6K2 and IP6K3, which have distinct biological functions. Of these, Inositol hexakisphosphate kinase‐2 (IP6K2), has been identified as a pro‐apoptotic factor with the ability to sensitize cells to apoptosis. To identify the protein interactome of IP6K2, we subjected wild‐type mouse brain tissue lysates to anti‐IP6K2 antibody‐mediated coimmuno‐precipitation followed by Liquid Chromatography Tandem Mass‐Spectrometry (LC‐MS/MS). This screen revealed robust binding of protein 4.1N with IP6K2. 4.1N is believed to confer stability and plasticity to the neuronal membrane via interactions with multiple binding partners, including the spectrin‐actin‐based cytoskeleton, integral membrane channels and receptors as well as membrane‐associated guanylate kinases. To further assess the specificity of the observed interaction between IP6K2 and 4.1N, we separately immuno‐precipitated IP6K1 and IP6K3 from brain tissue lysates of mice and failed to detect any association with the 4.1N protein. Thus, the interaction of 4.1N is evidently specific to the IP6K2 form of inositol hexakisphosphate kinases. Moreover, nuclear translocation of 4.1N, which is required for its principal functions, was dependent on IP6K2. We also found that IP6K2 and 4.1N are expressed in the granule cells and that their interaction regulate Purkinje cell morphology and synapse in the cerebellum. This was determined through Golgi staining, double immunofluorescence staining and electron microscopy. Deletion of IP6K2 in mice led to substantial defects in synaptic influences of granule cells upon Purkinje cells. Rotarod, open field test and Gait analysis also revealed the defects of motor coordination in IP6K2 knockout mice indicating the functional consequences of these interaction. Overall, our study establishes the functional implication of the robust association between IP6K2‐4.1N in the brain. Support or Funding Information The present funding capabilities and strategies of our lab do not have provisions for covering the defined expenditure related to the current scientific meeting. Thus, to enrich my research skills and knowledge in neurophysiology as well as inositol phosphates, I would request possible financial support through the Postdoctoral Travel Award funds to help me attend this meeting. This would allow me to share my work and ideas with young scientists as well as researchers from US and other parts of the world thus helping me to get my research ideas as well as work, evaluated and advised on, by the best in the world I would therefore entreat your good offices to consider my present submission for possible financial aid to help me attend the present meeting on Experimental Biology 2018 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .