Transcriptomic effects of Htt intron 1 polyA site deletion in HdhQ150 mice at 6 months

We have used a CRISPR-Cas9 approach to prevent the production of HTT1a in a knock-in mouse models of Huntington’s disease. All potential cryptic polyA sites were deleted from Htt intron 1 in HdhQ150 mice and colonies were established that were heterozygous for the intron 1 deletion on a mutant allele (HdhQ150ΔI) and heterozygous for the deletion on a wild-type allele (WTΔI). The CAG repeat sizes in the HdhQ150 and HdhQ150ΔI colonies were well-matched at approximately 190 CAGs.

As predicted, the deletion of Htt intron 1 cryptic polyA sites prevented termination of transcription in intron 1, resulting in the ablation of Htt1a in the HdhQ150ΔI mice. However, there were very low levels of the HTT1a protein, which resulted from a Htt readthrough product, that had retained the deleted intron and terminated at a cryptic polyA site in intron 2. HdhQ150, HdhQ150ΔI, wild-type and WTΔI mice were studied until 17 months of age.

Immunohistochemical and homogeneous time resolved fluorescence analysis showed that HTT aggregation in both HdhQ150 and HdhQ150ΔI brains contained HTT1a, but the dramatic decrease in soluble HTT1a levels in HdhQ150ΔI brains delayed the appearance of aggregated HTT1a by several months. Although this delay in aggregate pathology only partially reversed transcriptional dysregulation, the biomarkers NEFL and BRP39 (YKL40) remained at wild-type levels in HdhQ150ΔI mice at 17 months of age.

These data demonstrate that the production of HTT1a initiates HTT aggregation and that it is important to target HTT1a in huntingtin-lowering therapeutic strategies.