Mouse models for hereditary spastic paraplegia uncover a role of PI4K2A in autophagic lysosome reformation.

ABSTRACT

Hereditary spastic paraplegia (HSP) denotes genetically heterogeneous disorders characterized by leg spasticity due to degeneration of corticospinal axons. SPG11 and SPG15 have a similar clinical course and together are the most prevalent autosomal recessive HSP entity. The respective proteins play a role for macroautophagy/autophagy and autophagic lysosome reformation (ALR). Here, we report that spg11 and zfyve26 KO mice developed motor impairments within the same course of time. This correlated with enhanced accumulation of autofluorescent material in neurons and progressive neuron loss. In agreement with defective ALR, tubulation events were diminished in starved KO mouse embryonic fibroblasts (MEFs) and lysosomes decreased in neurons of KO brain sections. Confirming that both proteins act in the same molecular pathway, the pathologies were not aggravated upon simultaneous disruption of both. We further show that PI4K2A (phosphatidylinositol 4-kinase type 2 alpha), which phosphorylates phosphatidylinositol to phosphatidylinositol-4-phosphate (PtdIns4P), accumulated in autofluorescent deposits isolated from KO but not WT brains. Elevated PI4K2A abundance was already found at autolysosomes of neurons of presymptomatic KO mice. Immunolabelings further suggested higher levels of PtdIns4P at LAMP1-positive structures in starved KO MEFs. An increased association with LAMP1-positive structures was also observed for clathrin and DNM2/dynamin 2, which are important effectors of ALR recruited by phospholipids. Because PI4K2A overexpression impaired ALR, while its knockdown increased tubulation, we conclude that PI4K2A modulates phosphoinositide levels at autolysosomes and thus the recruitment of downstream effectors of ALR. Therefore, PI4K2A may play an important role in the pathogenesis of SPG11 and SPG15.Abbreviations ALR autophagic lysosome reformation; AP-5 adaptor protein complex 5; BFP blue fluorescent protein; dKO double knockout; EBSS Earle's balanced salt solution; FBA foot base angle; GFP green fluorescent protein; HSP hereditary spastic paraplegia; KO knockout; LAMP1 lysosomal-associated membrane protein 1; MAP1LC3B/LC3 microtubule-associated protein 1 light chain 3 beta; MEF mouse embryonic fibroblast; SQSTM1/p62 sequestosome 1; PI4K2A phosphatidylinositol 4-kinase type 2 alpha; PtdIns3P phosphatidylinositol-3-phosphate; PtdIns4P phosphatidylinositol-4-phosphate; RFP red fluorescent protein; SPG spastic paraplegia gene; TGN trans-Golgi network; WT wild type.