Intermediate filament aggregation in fibroblasts of giant axonal neuropathy patients is aggravated in non dividing cells and by microtubule destabilization

Abstract

Giant axonal neuropathy (GAN) is a severe neurodegenerative disorder characterized by the accumulation of neurofilaments (NFs) in distended axons. GAN corresponds to a disorder of the cytoplasmic intermediate filaments (IFs), since an abnormal aggregation of different IFs has been reported in several cell types, including NFs in neurons and vimentin in fibroblasts. The recent identification of the defective protein, named gigaxonin, now renders possible investigations on the mechanisms that trigger the destabilization of IFs. Although gigaxonin domain organization suggests multiple protein-protein interactions, via the BTB and the Kelch domains, the low amino acid identity with other members of the BTB/Kelch subfamily did not allow hypothesis about its function. In the present work, we studied GAN primary fibroblasts, and show that vimentin aggregation suffers great variation on prolonged culture at confluence and in low serum condition. While neither the microfilament (MF) nor the microtubule (MT) networks are perturbed by vimentin destabilization, we found that the aggregates are in close proximity to the microtubule organizing centers (MTOCs). Moreover, we show that MTs depolymerization induces a total vimentin aggregation in GAN fibroblasts. The results, together with the recent finding of an interaction between gigaxonin and MAP1B, a MT associated protein, suggests that gigaxonin plays an important role in the crosstalk between the IF and MT networks. We found that, when overexpressed, gigaxonin is localized in the cytoplasm but does not colocalize with any of the cytoskeletal networks, suggesting that the presence of the binding partner is rate limiting for proper localization of gigaxonin.