Disruption of the Autophagy-Lysosome Pathway Is Involved in Neuropathology of the nclf Mouse Model of Neuronal Ceroid Lipofuscinosis

Abstract

Variant late-infantile neuronal ceroid lipofuscinosis, a fatal lysosomal storage disorder accompanied by regional atrophy and pronounced neuron loss in the brain, is caused by mutations in the CLN6 gene. CLN6 is a non-glycosylated endoplasmic reticulum (ER)-resident membrane protein of unknown function. To investigate mechanisms contributing to neurodegeneration in CLN6 disease we examined the nclf mouse, a naturally occurring model of the human CLN6 disease. Prominent autofluorescent and electron-dense lysosomal storage material was found in cerebellar Purkinje cells, thalamus, hippocampus, olfactory bulb and in cortical layer II to V. Another prominent early feature of nclf pathogenesis was the localized astrocytosis that was evident in many brain regions and the more widespread microgliosis. Expression analysis of mutant Cln6 found in nclf mice demonstrated synthesis of a truncated protein with a reduced half-life. Whereas the rapid degradation of the mutant Cln6 protein can be inhibited by proteasomal inhibitors, there was no evidence for ER stress or activation of the unfolded protein response in various brain areas during postnatal development. Age-dependent increases in LC3-II, ubiquitinated proteins, and neuronal p62-positive aggregates were observed, indicating a disruption of the autophagy-lysosome degradation pathway of proteins in brains of nclf mice, most likely due to defective fusion between autophagosomes and lysosomes. These data suggest that proteasomal degradation of mutant Cln6 is sufficient to prevent the accumulation of misfolded Cln6 protein, whereas lysosomal dysfunction impairs constitutive autophagy promoting neurodegeneration.

Induction of Lysosomal Biogenesis in Atherosclerotic Macrophages Can Rescue Lipid-Induced Lysosomal Dysfunction and Downstream Sequelae

“…In the absence of an intact lysosomal machinery, aged and misfolded proteins aggregate in cytoplasmic precipitates that can be toxic to the cell 29 . In support of this, treatment of peritoneal macrophages with atherogenic lipids (oxLDL and cholesterol crystals) can increase inclusion body formation (Figure 6D, Control) likely by disrupting lysosomal function.…”

The Endolysosomal System in Cell Death and Survival

“…It was thus shown that lysosomal dysfunction promotes neurodegeneration because it impairs constitutive autophagy (Thelen et al 2012). In addition, it was proposed that the protective autophagic response induced by TRAIL continuously sequesters caspase-8 in autophagosomes, but its elimination depends on the subsequent lysosomal degradation (Hou et al 2010).…”

A Novel Interaction Between Aging and ER Overload in a Protein Conformational Dementia

“…Western blotting was performed under denaturing and nondenaturing (Miranda et al 2004) conditions using antibodies against GFP (Clonetch), a-tubulin (Sigma-Aldrich), ATF6 (Imgenex), eIF2a/eIF2aP (Cell Signaling), neuroserpin (Miranda et al 2008), LC3-I/-II (Thelen et al 2012), and actin (Sigma-Aldrich). Visualization and quantification was done with either Imager Gel Doc System and Quantity One software (Bio-Rad) or Typhoon Trio scanner with ImageQuant software (GE Healthcare).…”

“…Visualization and quantification was done with either Imager Gel Doc System and Quantity One software (Bio-Rad) or Typhoon Trio scanner with ImageQuant software (GE Healthcare). Changes in protein expression of ATF6, LC3-I, LC3-II, or eIF2aP were calculated by normalization to b-actin or eI2Fa in three independent experiments, where wild-type values were set to one (Glatzel et al 2001;Thelen et al 2012).…”