Intrinsic mutant HTT-mediated defects in oligodendroglia cause myelination deficits and behavioral abnormalities in Huntington disease

Abstract

White matter abnormalities are a nearly universal pathological feature of neurodegenerative disorders including Huntington disease (HD). A long-held assumption is that this white matter pathology is simply a secondary outcome of the progressive neuronal loss that manifests with advancing disease. Using a mouse model of HD, here we show that white matter and myelination abnormalities are an early disease feature appearing before the manifestation of any behavioral abnormalities or neuronal loss. We further show that selective inactivation of mutant huntingtin (mHTT) in the NG2+ oligodendrocyte progenitor cell population prevented myelin abnormalities and certain behavioral deficits in HD mice. Strikingly, the improvements in behavioral outcomes were seen despite the continued expression of mHTT in nonoligodendroglial cells including neurons, astrocytes, and microglia. Using RNA-seq and ChIP-seq analyses, we implicate a pathogenic mechanism that involves enhancement of polycomb repressive complex 2 (PRC2) activity by mHTT in the intrinsic oligodendroglial dysfunction and myelination deficits observed in HD. Our findings challenge the long-held dogma regarding the etiology of white matter pathology in HD and highlight the contribution of epigenetic mechanisms to the observed intrinsic oligodendroglial dysfunction. Our results further suggest that ameliorating white matter pathology and oligodendroglial dysfunction may be beneficial for HD.

Keywords: Huntington disease; PRC2; myelination; oligodendrocytes; white matter.

Fig. 1.

OPC-intrinsic effects of mHTT cause myelination abnormalities in HD mice. (A) Schematic representation of Cre-mediated genetic reduction of mHTT expression in OPCs (NG2+ cells) in BACHD mice. (B) PCR analysis confirmed the excision of human mHTT exon 1 in the cortex of BN mice. (C) mHTT mRNA levels are reduced in purified OPCs in BN mice at P6–P7. n = 3/genotype (P = 0.0100, t = 4.601, df = 4). (D) EM images of myelinated axons in the CC at 12 mo of age. (Scale bar, 1 μm.) (E–G) Higher g-ratios (thinner myelin sheaths) in BACHD mice are rescued in BN mice. n = 3/genotype; ∼300 axons were quantified per animal. Data show means ± SEM; *P < 0.05, **P < 0.01; two-tailed Student’s test in C and one-way ANOVA followed by Tukey’s test in G.

Fig. 2.

Behavioral deficits in HD mice are partly the result of mHTT-mediated defects in oligodendroglia. (A) Overview of behavioral assessments. BACHD mice show cognitive deficits in the rotarod learning (B), motor deficits in the rotarod (C) and climbing (D) tests, anxiety-like behavior in the OF at 6 mo of age (E), and depressive-like behavior in the Porsolt FST at 12 mo of age (F). The ability to swim is comparable among genotypes at 12 mo of age (G). BN mice show a rescue in some of the behavioral phenotypes. n = 12–20 mixed gender/genotype. Data represent means ± SEM; ns, not significant; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 (compared with BACHD); ^###P < 0.001; ^####P < 0.0001 (compare with BN). One-way ANOVA (E–G) or two-way ANOVA (B–D) followed by Tukey’s multiple comparisons test were applied for all behavioral studies. A, age; G, genotype; T, trial.

Fig. 3.

Epigenetic dysregulation mediates mHTT effects on oligodendroglia. (A) Heatmap and hierarchal clustering of the significantly differentially expressed genes between WT (n = 3), BACHD (n = 3), and BN (n = 3) (360 genes, 10% FDR likelihood ratio test). Red indicates higher gene expression, and blue represents lower gene expression. Boxes indicate clusters of samples determined by 10,000 bootstraps. (B) Volcano plot showing the differentially expressed genes between BN (n = 3) and BACHD (n = 3) mouse corpus callosum. The significant up-regulated genes with respect to BN are indicated in red, while the significant down-regulated genes are indicated in blue (FDR < 10%). (C) Gene ontology analysis of significant DEGs between BACHD and BN mice. The top three significant terms (FDR < 5%) for up-regulated and down-regulated genes are shown. (D) Heat map shows mean gene expression levels of selected genes in WT, BACHD, and BN mice. (E) Nkx2.2 appears as the top motif enriched in up-regulated DEGs between BACHD and BN. (F) Htt gene expression (fragments per kilobase million) in different stages of oligodendroglial differentiation (data from ref. ; n = 2 for each group, and bars indicate mean). MOL, myelinating oligodendrocytes; NFOL, newly formed oligodendrocytes; OPC, oligodendrocyte progenitor cells. (G) REST and PRC2-binding sites are enriched in DEGs between BACHD and BN. (H) ChIP-qPCR enrichment at the En2 promoter in CC for EZH2 and SUZ12. Rpl32 was used as negative control. (I) Increased number of EZH2- and SUZ12-binding sites in the BACHD mice compared with WT is partially rescued in BN mice. (J) Immunoblot analysis of H3K27me3 in the CC of WT, BACHD, and BN mice. Values normalized to WT and presented as means ± SEM; n = 3 per genotype; *P < 0.05 by one-way ANOVA with Tukey’s post hoc test; ^$P < 0.05 by unpaired two-tailed t test.