Genetic Deletion of Thorase Causes Purkinje Cell Loss and Impaired Motor Coordination Behavior.

Thorase belongs to the AAA+ ATPase family, which plays a critical role in maintaining cellular homeostasis. Our previous work reported that Thorase was highly expressed in brain tissue, especially in the cerebellum. However, the roles of Thorase in the cerebellum have still not been characterized. In this study, we generated conditional knockout mice (cKO) with Thorase deletion in Purkinje cells. Thorase cKO mice exhibited cerebellar degenerative diseases-like behavior and significant impairment in motor coordination. Thorase deletion resulted in more Purkinje neuron apoptosis, leading to Purkinje cell loss in the cerebellum of Thorase cKO mice. We also found enhanced expression of the inflammatory protein ASC, IL-1ß, IL-6 and TNF-α in the Thorase cKO cerebellum, which contributed to the pathogenesis of cerebellar degenerative disease. Our findings provide a better understanding of the role of Thorase in the cerebellum, which is a theoretical basis for Thorase as a therapeutic drug target for neurodegenerative diseases.

ATPase Thorase Deficiency Causes α-Synucleinopathy and Parkinson's Disease-like Behavior.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases and is pathologically characterized by α-synucleinopathy, which is harmful to dopaminergic neurons. However, the underlying mechanisms and pathogenesis of PD remain unclear. The AAA + ATPase Thorase was identified as being essential for neuroprotection and synaptic plasticity by regulating the AMPA receptor trafficking. Here, we found that conditional knockout of Thorase resulted in motor behaviors indicative of neurodegeneration. Genetic deletion of Thorase exacerbated phenotypes of α-synucleinopathy in a familial PD-like A53T mouse model, whereas overexpression of Thorase prevented α-syn accumulation in vivo. Biochemical and cell cultures studies presented here suggest that Thorase interacts with α-syn and regulates the degradation of ubiquitinated α-syn. Thorase deficiency promotes α-syn aggregation in primary cultured neurons. The discoveries in this study provide us with a further understanding of the pathogenesis of α-synucleinopathies including PD.

Transmembrane dislocases: a second chance for protein targeting.

Precise distribution of proteins is essential to sustain the viability of cells. A complex network of protein synthesis and targeting factors cooperate with protein quality control systems to ensure protein homeostasis. Defective proteins are inevitably degraded by the ubiquitin-proteasome system and lysosomes. However, due to overlapping targeting information and limited targeting fidelity, certain proteins become mislocalized. In this review, we present the idea that transmembrane dislocases recognize and remove mislocalized membrane proteins from cellular organelles. This enables other targeting attempts and prevents degradation of mislocalized but otherwise functional proteins. These transmembrane dislocases can be found in the outer mitochondrial membrane (OMM) and endoplasmic reticulum (ER). We highlight common principles regarding client recognition and outline open questions in our understanding of transmembrane dislocases.

AMPA Receptor Surface Expression Is Regulated by S-Nitrosylation of Thorase and Transnitrosylation of NSF.

The regulation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) trafficking affects multiple brain functions, such as learning and memory. We have previously shown that Thorase plays an important role in the internalization of AMPARs from the synaptic membrane. Here, we show that N-methyl-d-aspartate receptor (NMDAR) activation leads to increased S-nitrosylation of Thorase and N-ethylmaleimide-sensitive factor (NSF). S-nitrosylation of Thorase stabilizes Thorase-AMPAR complexes and enhances the internalization of AMPAR and interaction with protein-interacting C kinase 1 (PICK1). S-nitrosylated NSF is dependent on the S-nitrosylation of Thorase via trans-nitrosylation, which modulates the surface insertion of AMPARs. In the presence of the S-nitrosylation-deficient C137L Thorase mutant, AMPAR trafficking, long-term potentiation, and long-term depression are impaired. Overall, our data suggest that both S-nitrosylation and interactions of Thorase and NSF/PICK1 are required to modulate AMPAR-mediated synaptic plasticity. This study provides critical information that elucidates the mechanism underlying Thorase and NSF-mediated trafficking of AMPAR complexes.

Ganglioside regulation of AMPA receptor trafficking.

Gangliosides are major cell-surface determinants on all vertebrate neurons. Human congenital disorders of ganglioside biosynthesis invariably result in intellectual disability and are often associated with intractable seizures. To probe the mechanisms of ganglioside functions, affinity-captured ganglioside-binding proteins from rat cerebellar granule neurons were identified by quantitative proteomic mass spectrometry. Of the six proteins that bound selectively to the major brain ganglioside GT1b (GT1b:GM1 > 4; p < 10(-4)), three regulate neurotransmitter receptor trafficking: Thorase (ATPase family AAA domain-containing protein 1), soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (γ-SNAP), and the transmembrane protein Nicalin. Thorase facilitates endocytosis of GluR2 subunit-containing AMPA-type glutamate receptors (AMPARs) in an ATPase-dependent manner; its deletion in mice results in learning and memory deficits (J. Zhang et al., 2011b). GluR2-containing AMPARs did not bind GT1b, but bound specifically to another ganglioside, GM1. Addition of noncleavable ATP (ATPγS) significantly disrupted ganglioside binding, whereas it enhanced AMPAR association with Thorase, NSF, and Nicalin. Mutant mice lacking GT1b expressed markedly higher brain Thorase, whereas Thorase-null mice expressed higher GT1b. Treatment of cultured hippocampal neurons with sialidase, which cleaves GT1b (and other sialoglycans), resulted in a significant reduction in the size of surface GluR2 puncta. These data support a model in which GM1-bound GluR2-containing AMPARs are functionally segregated from GT1b-bound AMPAR-trafficking complexes. Release of ganglioside binding may enhance GluR2-containing AMPAR association with its trafficking complexes, increasing endocytosis. Disrupting ganglioside biosynthesis may result in reduced synaptic expression of GluR2-contianing AMPARs resulting in intellectual deficits and seizure susceptibility in mice and humans.