Cytosolic PSD-95 interactor (cypin) is a multifunctional, guanine deaminase that plays a major role in shaping the morphology of the dendritic arbor of hippocampal and cortical neurons. Cypin catalyzes the Zn2+-dependent deamination of guanine to xanthine, which is then metabolized to uric acid by xanthine oxidase. Cypin binds to tubulin heterodimers via its carboxyl terminal region (amino acids (aa) 350-454), which contains a collapsin response mediator protein (CRMP) homology domain (aa 350-403). Moreover, this region alone is not sufficient to facilitate microtubule polymerization; therefore, additional cypin regions must be involved in this process. Here, we asked whether cypin binds to fully formed microtubules and how overexpression of cypin regulates the microtubule cytoskeleton in dendrites of cultured hippocampal neurons. Protein-protein docking strategies confirm that the cypin homodimer binds to tubulin heterodimers via amino acids within aa 350-454. Biochemical pull-down data suggest that aa 1-220 are necessary for cypin binding to soluble tubulin heterodimers and to taxol-stabilized microtubules. Molecular docking of the cypin homodimer to soluble tubulin heterodimers reveals a consistently observed docking pose using aa 47-71, 113-118, 174-178, and 411-418, which is consistent with our biochemical data. Additionally, overexpression of cypin in hippocampal neurons results in decreased spacing between microtubules. Our results suggest that several protein domains facilitate cypin-mediated polymerization of tubulin heterodimers into microtubules, possibly through a mechanism whereby cypin dimers bind to multiple tubulin heterodimers.
Keywords: Cytosolic PSD-95 interactor; Hippocampal neuron; Microtubule polymerization; Microtubule protofilament; Tubulin heterodimer.
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