Abstract
Microtubules play a critical role in multiple aspects of neurodevelopment, including the generation, migration and differentiation of neurons. A recurrent mutation (R402H) in the α-tubulin gene TUBA1A is known to cause lissencephaly with cerebellar and striatal phenotypes. Previous work has shown that this mutation does not perturb the chaperone-mediated folding of tubulin heterodimers, which are able to assemble and incorporate into the microtubule lattice. To explore the molecular mechanisms that cause the disease state we generated a new conditional mouse line that recapitulates the R402H variant. We show that heterozygous mutants present with laminar phenotypes in the cortex and hippocampus, as well as a reduction in striatal size and cerebellar abnormalities. We demonstrate that homozygous expression of the R402H allele causes neuronal death and exacerbates a cell intrinsic defect in cortical neuronal migration. Microtubule sedimentation assays coupled with quantitative mass spectrometry demonstrated that the binding and/or levels of multiple microtubule associated proteins (MAPs) are perturbed by the R402H mutation including VAPB, REEP1, EZRIN, PRNP and DYNC1l1/2. Consistent with these data we show that the R402H mutation impairs dynein-mediated transport which is associated with a decoupling of the nucleus to the microtubule organising center. Our data support a model whereby the R402H variant is able to fold and incorporate into microtubules, but acts as a gain of function by perturbing the binding of MAPs.
Highlights
Consisting of α and β tubulin heterodimers, microtubules are dynamic polymers that play a critical role in multiple aspects of neuronal development, including the generation, migration and differentiation of neurons [1,2]
Microtubules are polymers composed of tubulin proteins, which play an important role in the development of the human brain
We have previously shown that the R402H mutation does not perturb the chaperonemediated folding of tubulin heterodimers, and that R402H mutant heterodimers can integrate into the interphase cytoskeleton [16]
Summary
Consisting of α and β tubulin heterodimers, microtubules are dynamic polymers that play a critical role in multiple aspects of neuronal development, including the generation, migration and differentiation of neurons [1,2]. As neurons exit from the proliferative ventricular zone they transition from a multipolar to a bipolar morphology, migrating along radial glia into the developing cortical plate [3]. This process requires an array of molecules that are responsible for a dramatic reorganisation of the cell’s cytoskeletal architecture, and the generation of force to translocate the nucleus [4]. Doublecortin (DCX) is a microtubuleassociated protein and LIS1 plays an important role in regulating dynein, a cytoskeletal molecular motor [5,6]. Mutations in DCX and LIS1 genes have been shown to cause lissencephaly, whilst dynein variants are associated with a range of cortical malformations [7,8,9]
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