Department of Cell and Developmental Biology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.
Ken and Ruth Davee Department of Neurology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.
FASEB J. 2023 May;37(5):e22886. doi: 10.1096/fj.202202119R.
Gigaxonin is an adaptor protein for E3 ubiquitin ligase substrates. It is necessary for ubiquitination and degradation of intermediate filament (IF) proteins. Giant axonal neuropathy is a pathological condition caused by mutations in the GAN gene that encodes gigaxonin. This condition is characterized by abnormal accumulation of IFs in both neuronal and non-neuronal cells; however, it is unclear what causes IF aggregation. In this work, we studied the dynamics of IFs using their subunits tagged with a photoconvertible protein mEOS 3.2. We have demonstrated that the loss of gigaxonin dramatically inhibited transport of IFs along microtubules by the microtubule motor kinesin-1. This inhibition was specific for IFs, as other kinesin-1 cargoes, with the exception of mitochondria, were transported normally. Abnormal distribution of IFs in the cytoplasm can be rescued by direct binding of kinesin-1 to IFs, demonstrating that transport inhibition is the primary cause for the abnormal IF distribution. Another effect of gigaxonin loss was a more than 20-fold increase in the amount of soluble vimentin oligomers in the cytosol of gigaxonin knock-out cells. We speculate that these oligomers saturate a yet unidentified adapter that is required for kinesin-1 binding to IFs, which might inhibit IF transport along microtubules causing their abnormal accumulation.
巨轴索神经病是一种由编码 gigaxonin 的 GAN 基因突变引起的病理状况。这种情况的特征是中间丝(IF)在神经元和非神经元细胞中的异常积累;然而,尚不清楚是什么导致 IF 聚集。在这项工作中,我们使用标记有光可转化蛋白 mEOS 3.2 的 IF 亚基研究了 IF 的动力学。我们已经证明,gigaxonin 的缺失严重抑制了微管动力蛋白 kinesin-1 沿微管的 IF 运输。这种抑制是 IF 特有的,因为除了线粒体之外,其他 kinesin-1 货物都能正常运输。IF 在细胞质中的异常分布可以通过 kinesin-1 与 IF 的直接结合来挽救,这表明运输抑制是 IF 异常分布的主要原因。gigaxonin 缺失的另一个影响是细胞溶质中可溶性波形蛋白寡聚物的含量增加了 20 多倍在 gigaxonin 敲除细胞中。我们推测这些寡聚物饱和了一种尚未鉴定的衔接蛋白,该蛋白是 kinesin-1 与 IF 结合所必需的,这可能会抑制 IF 沿微管的运输,导致其异常积累。