Enrichment in stable microtubules reveals dysfunction of microtubule dynamics occurring in Parkinson’s disease models

Microtubules (MTs) are dynamically unstable polymers that interconvert stochastically between polymerization and depolymerization phases, a behavior known as dynamic instability. During neuronal differentiation MTs are highly dynamic to promote process outgrowth, whereas in mature neurons their stability increase to ensure cell structure, function and viability. However, proper microtubule dynamics remains fundamental for the survival of the mature neuron, and has a narrow range of activity. If microtubule dynamics overcomes these limits, microtubule-dependent function are compromised and cell death occurs. In this regard, much attention has been placed on the link between microtubule dysfunction and neurodegeneration (Feinstein and Wilson, 2005). Focusing on Parkinson’s disease (PD), recent data indicate that alpha-synuclein and parkin, two protein mutated in PD, interact with MTs and seem to stabilize them; moreover, the parkinsonism-inducing neurotoxin rotenone and MPP+, toxic metabolite of MPTP, are able to destabilize MTs both in vitro and in cellular model of PD. We reported earlier that that MPP+ causes an increase of the unpolymerized fraction in cultured cells and acts as catastrophe inductor in MTs assembled from purified tubulin (Cappelletti et al. 1999, 2005). Here we address the question of stability of MTs in cellular and animal models of PD. On the ground that novel and highly dynamic MTs are enriched in tyrosinated (Tyr) tubulin whereas long-lived and stable microtubules are enriched in detyrosinated (Glu) tubulin (Westermann and Weber, 2003), we evaluated the levels of Glu-tubulin in extracts obtained from MPP+-treated PC12 cells and MPTP-treated mice. We found increased levels of Glu-tubulin both in cultured cells and in the corpus striatum of mice following the treatment. These data show that the neurotoxin induces the increase of stable subset of MTs and suggest a reduced MT dynamics. In order to get further insight into the effect of MPP+ on MTs dynamics, we followed the growing end of MTs in PC12 cells transfected with EB3-GFP protein by live cell imaging microscopy. We found that MPP+ affects the dynamics of MTs and causes the overall slowing down in their growth. Our results are consistent with previous data on reconstituted system of MTs and strengthen the central role of MTs in the pathogenesis of PD.