A natural antisense transcript at the Huntington's disease repeat locus regulates HTT expression

Abstract

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a CAG repeat expansion in exon 1 of huntingtin (HTT). Relatively little attention has been directed to the genomic features of the antisense strand at the HD locus, though the presence of a transcript from this strand has been suggested by a survey of the entire transcriptome and the existence of several EST tags. In this study, we identified huntingtin antisense (HTTAS), a natural antisense transcript at the HD repeat locus that contain the repeat tract. HTTAS is 5' capped, poly (A) tailed and contains three exons, alternatively spliced into HTTAS_v1 (exons 1 and 3) and HTTAS_v2 (exons 2 and 3). Exon 1 includes the repeat. HTTAS_v1 has a weak promoter, and is expressed at low levels in multiple tissue types and throughout the brain. Reporter assays indicate that while efficient promoter activity requires a short repeat, repeat expansion reduces promoter efficiency. Consistent with the reporter assays, levels of HTTAS_v1 are reduced in human HD frontal cortex. In cell systems, overexpression of HTTAS_v1 specifically reduces endogenous HTT transcript levels, while siRNA knockdown of HTTAS_v1 increases HTT transcript levels. Minigene constructs of the HD locus confirm the regulatory effect of HTTAS_v1 on HTT, and demonstrate that the effect is dependent on repeat length and is at least partially Dicer dependent. Together, these findings provide strong evidence for the existence of a gene antisense to HTT, with properties that include regulation of HTT expression.