The unstructured N-terminal region of doublecortin X (DCX) makes critical contributions to microtubule organization

M Moslehi1,2, DCH Ng3 and MA Bogoyevitch1,2

  1. Regulatory Biology Group, Cell Signalling Research Laboratories, School of Biomedical Sciences, University of Melbourne, Parkville, VIC 3010
  2. Regulatory Biology Group, Department of Biochemistry, Bio21 Institute, University of Melbourne, Parkville, VIC 3052
  3. Cell Signalling Laboratory, School of Biomedical Sciences, University of Queensland, St Lucia, QLD 4072

Doublecortin X (DCX) is a neuron-specific microtubule-associated protein that is essential for the cortical layering and neuronal migration in developing brain. Although DCX directly interacts with microtubules via its two structured doublecortin (DC) domains, the unstructured flanking N- and C-terminal regions of DCX have been proposed to play putative regulatory roles albeit by largely unknown mechanisms. We are investigating the contributions made by the unstructured DCX N-terminus (DCX 1-45) towards ability of DCX to organize microtubules and to regulate microtubule-related events. Whilst wild-type DCX expression leads to pronounced microtubule reorganisation into microtubule bundles in both neuronal and non-neuronal cell types, a progressive deletion analysis has revealed that loss of the first 29 amino acids was sufficient to prevent this impact on the microtubules. Furthermore, a DCX mutant lacking the entire N-terminus DCX-ΔN1-45 was unable to interact with microtubules as demonstrated in in vivo microtubule-binding assays. Expression of this DCX-ΔN1-45 mutant also impeded microtubule regrowth following nocodazole-induced microtubule depolymerisation. In parallel, we are also evaluating the contributions of phosphorylation of the N-terminus to regulation of DCX by the expression of phospho-mimetic or non-phosphorylated DCX mutants, again demonstrating an impact of these different DCX mutants on the DCX dynamics with microtubules as well as microtubule-bundling. Taken together, these results emphasize the critical regulation of DCX-microtubule interaction by DCX N-terminus and future work will tackle how these biochemical events contribute to regulation of neuronal migration.