Mechanisms of Translational Dysregulation in Huntington Disease - This investigation aimed to identify the mechanisms and molecular functions of ribosome stalling in Huntington disease (HD). HD is caused by the polyglutamine (CAG) expansion of huntingtin (mHTT), which promotes neurodegeneration in the brain, causing motor, cognitive, and psychiatric symptoms. Multiple abnormal functions have been proposed for mHTT, but its pathogenic mechanisms remain unclear. Using super-resolution ribosome profiling (aka Ribo-Seq) and biochemical tools, we demonstrate that mHTT promotes ribosome stalling and inhibits protein synthesis, which is the topic of the current investigation. The central hypothesis is that mHTT interacts with RNA and ribosomal binding proteins (RBPs) to inhibit ribosomal movement and increase HD pathogenesis. Our published and preliminary data indicate that mHTT copurifies with 40S ribosomal subunits, directly binds to elongating ribosomes, and suppresses protein synthesis both in vitro and in vivo. Interestingly, Ribo-Seq revealed that mHTT mRNA exon1 also had substantial ribosome occupancy before CAG expanded repeats, which we validated using biochemical reporter experiments. We found that the RBP fragile x-mental retardation protein (Fmrp), a negative regulator of synaptic mRNA translation, is upregulated in HD and that Fmrp deletion inhibits age-dependent motor deficits in a humanized HD model. On the basis of these solid data, the proposed research has three interrelated objectives: 1) to identify the ribosome-bound protein/RNA component that interacts with mHTT; 2) to determine the mechanisms of ribosome stalling on mHTT mRNA; and 2) to delineate the role of Fmrp in modulating protein synthesis at the synaptic level in HD. Consequently, the outcomes will likely reveal novel hypothesis-generating ribosome stalling mechanisms in HD, identify critical cis and trans components implicated in mHTT RNA stalling, and determine how the mHTT-Fmrp nexus regulates aberrant synaptic translation in HD. These findings will help develop translation mechanism-based treatment and diagnostic modalities for poorly understood neurological illnesses.
