Huntingtin Expression Stimulates Endosomal–Lysosomal Activity, Endosome Tubulation, and Autophagy

Abstract

An expansion of polyglutamines in the N terminus of huntingtin causes Huntington's disease (HD) and results in the accrual of mutant protein in the nucleus and cytoplasm of affected neurons. How mutant huntingtin causes neurons to die is unclear, but some recent observations suggest that an autophagic process may occur. We showed previously that huntingtin markedly accumulates in endosomal-lysosomal organelles of affected HD neurons and, when exogenously expressed in clonal striatal neurons, huntingtin appears in cytoplasmic vacuoles causing cells to shrink. Here we show that the huntingtin-enriched cytoplasmic vacuoles formed in vitro internalized the lysosomal enzyme cathepsin D in proportion to the polyglutamine-length in huntingtin. Huntingtin-labeled vacuoles displayed the ultrastructural features of early and late autophagosomes (autolysosomes), had little or no overlap with ubiquitin, proteasome, and heat shock protein 70/heat shock cognate 70 immunoreactivities, and altered the arrangement of Golgi membranes, mitochondria, and nuclear membranes. Neurons with excess cytoplasmic huntingtin also exhibited increased tubulation of endosomal membranes. Exogenously expressed human full-length wild-type and mutant huntingtin codistributed with endogenous mouse huntingtin in soluble and membrane fractions, whereas human N-terminal huntingtin products were found only in membrane fractions that contained lysosomal organelles. We speculate that mutant huntingtin accumulation in HD activates the endosomal-lysosomal system, which contributes to huntingtin proteolysis and to an autophagic process of cell death.

Mutant Huntingtin Impairs Post-Golgi Trafficking to Lysosomes by Delocalizing Optineurin/Rab8 Complex from the Golgi Apparatus

“…These findings are coincident with the reduction of lysosomal activity observed in mhtt cells. Interestingly, we observed that overexpression of truncated forms of mhtt, such as exon1-103Q-htt, stimulated post-Golgi transport of Lamp-1 and also increased the labeling of cathepsin D in enlarged LysoTrackerpositive structures (Supplemental Figure 5), which is in agreement with a previous study (Kegel et al, 2000). This effect could be explained by several protein interactions that are lost in the case of shorter fragments of htt.…”

Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease

“…Polyglutamine expansion induces autophagy mTOR inhibition induces autophagy, which has been previously reported in brains of individuals with Huntington disease, in transgenic mice expressing mutant huntingtin fragments and in a cell model of Huntington disease, using morphological criteria 23,24 . It has been difficult to differentiate autophagosomes from other vesicles in the absence of specific autophagosome markers.…”

“…mTOR was diffusely distributed in untransfected cells, in cells expressing wild-type protein (Gln 23 ; Fig. 1a) and in cells expressing mutant protein (Gln 74 ) without aggregates but colocalized with both cytoplasmic and nuclear aggregates in >98% of mutant cells with aggregates ( Fig.…”

“…Furthermore, levels of LC3-II increased after expression of mutant huntingtin was induced in stable PC12 cells (data not shown), whereas no difference was seen upon induction of expression of wild-type huntingtin. Autophagic activity also correlates with the number of LC3-stained vesicles 25 , which increased after serum starvation or treatment with rapamycin (known to stimulate autophagy) and in aggregate-containing cells expressing mutant Gln 74 or Gln 103 huntingtin but not in cells expressing wild-type Gln 23 or Gln 25 huntingtin ( Fig. 4b and Supplementary Fig.…”

Role of the pro-survival molecule Bfl-1 in melanoma

“…Thus, it is advisable to take the term 'autophagic cell death' to mean cell death occurring with associated autophagy rather than as a result of autophagy. The role of autophagy in disease is not well understood; it appears to play a protective role in the progression of neurodegenerative diseases and muscular disorders and provide protection from infections [123][124][125] , but contributes to the pathology of other diseases such as…”