Huntingtin contains an ubiquitin-binding domain and regulates lysosomal targeting of mitochondrial and RNA-binding proteins.

Understanding the normal function of the Huntingtin (HTT) protein is of significance in the design and implementation of therapeutic strategies for Huntington's disease (HD). Expansion of the CAG repeat in the HTT gene, encoding an expanded polyglutamine (polyQ) repeat within the HTT protein, causes HD and may compromise HTT's normal activity contributing to HD pathology. Here, we investigated the previously defined role of HTT in autophagy specifically through studying HTT's association with ubiquitin. We find that HTT interacts directly with ubiquitin in vitro. Tandem affinity purification was used to identify ubiquitinated and ubiquitin-associated proteins that copurify with a HTT N-terminal fragment under basal conditions. Copurification is enhanced by HTT polyQ expansion and reduced by mimicking HTT serine 421 phosphorylation. The identified HTT-interacting proteins include RNA-binding proteins (RBPs) involved in mRNA translation, proteins enriched in stress granules, the nuclear proteome, the defective ribosomal products (DRiPs) proteome and the brain-derived autophagosomal proteome. To determine whether the proteins interacting with HTT are autophagic targets, HTT knockout (KO) cells and immunoprecipitation of lysosomes were used to investigate autophagy in the absence of HTT. HTT KO was associated with reduced abundance of mitochondrial proteins in the lysosome, indicating a potential compromise in basal mitophagy, and increased lysosomal abundance of RBPs which may result from compensatory up-regulation of starvation-induced macroautophagy. We suggest HTT is critical for appropriate basal clearance of mitochondrial proteins and RBPs, hence reduced HTT proteostatic function with mutation may contribute to the neuropathology of HD.

Transplanted human neural stem cells rescue phenotypes in zQ175 Huntington's disease mice and innervate the striatum.

Huntington's disease (HD), a genetic neurodegenerative disorder, primarily affects the striatum and cortex with progressive loss of medium-sized spiny neurons (MSNs) and pyramidal neurons, disrupting cortico-striatal circuitry. A promising regenerative therapeutic strategy of transplanting human neural stem cells (hNSCs) is challenged by the need for long-term functional integration. We previously described that, with short-term hNSC transplantation into the striatum of HD R6/2 mice, human cells differentiated into electrophysiologically active immature neurons, improving behavior and biochemical deficits. Here, we show that long-term (8 months) implantation of hNSCs into the striatum of HD zQ175 mice ameliorates behavioral deficits, increases brain-derived neurotrophic factor (BDNF) levels, and reduces mutant huntingtin (mHTT) accumulation. Patch clamp recordings, immunohistochemistry, single-nucleus RNA sequencing (RNA-seq), and electron microscopy demonstrate that hNSCs differentiate into diverse neuronal populations, including MSN- and interneuron-like cells, and form connections. Single-nucleus RNA-seq analysis also shows restoration of several mHTT-mediated transcriptional changes of endogenous striatal HD mouse cells. Remarkably, engrafted cells receive synaptic inputs, innervate host neurons, and improve membrane and synaptic properties. Overall, the findings support hNSC transplantation for further evaluation and clinical development for HD.

Treatment with JQ1, a BET bromodomain inhibitor, is selectively detrimental to R6/2 Huntington's disease mice.

Transcriptional and epigenetic alterations occur early in Huntington's disease (HD), and treatment with epigenetic modulators is beneficial in several HD animal models. The drug JQ1, which inhibits histone acetyl-lysine reader bromodomains, has shown promise for multiple cancers and neurodegenerative disease. We tested whether JQ1 could improve behavioral phenotypes in the R6/2 mouse model of HD and modulate HD-associated changes in transcription and epigenomics. R6/2 and non-transgenic (NT) mice were treated with JQ1 daily from 5 to 11 weeks of age and behavioral phenotypes evaluated over this period. Following the trial, cortex and striatum were isolated and subjected to mRNA-seq and ChIP-seq for the histone marks H3K4me3 and H3K27ac. Initially, JQ1 enhanced motor performance in NT mice. In R6/2 mice, however, JQ1 had no effect on rotarod or grip strength but exacerbated weight loss and worsened performance on the pole test. JQ1-induced gene expression changes in NT mice were distinct from those in R6/2 and primarily involved protein translation and bioenergetics pathways. Dysregulation of HD-related pathways in striatum was exacerbated by JQ1 in R6/2 mice, but not in NTs, and JQ1 caused a corresponding increase in the formation of a mutant huntingtin protein-dependent high molecular weight species associated with pathogenesis. This study suggests that drugs predicted to be beneficial based on their mode of action and effects in wild-type or in other neurodegenerative disease models may have an altered impact in the HD context. These observations have important implications in the development of epigenetic modulators as therapies for HD.