Cell-Type-Specific RNA Analysis Probes Selective Vulnerability in Huntington’s

How does mutant huntingtin protein cause striatal neurons to die? On July 17 in Neuron online, researchers led by Myriam Heiman at the Massachusetts Institute of Technology suggest that an early insult to mitochondria could kick off inflammation and neurodegeneration. Using multiple overlapping methods to catalogue gene-expression changes in striatal neurons of several mouse models and in Huntington’s postmortem brain tissue, the scientists found one of the earliest deficits to be a slowdown in mitochondrial oxidative phosphorylation. Mitochondrial RNA spilled out into cytoplasm, triggering an interferon-based inflammatory response from within the neurons. This was most pronounced in the spiny projection neurons primarily affected in Huntington’s disease, perhaps explaining the heightened vulnerability of these cells.

  • In Huntington’s striatal neurons, oxidative phosphorylation slows.
  • Mitochondrial RNA escapes into cytoplasm.
  • This activates an innate immune response that may lead to degeneration.

“The activation of innate immune signaling in the most vulnerable HD neurons provides a novel framework to understand the basis of mutant huntingtin toxicity, and raises new therapeutic opportunities,” the authors wrote.

To examine gene expression in specific neuronal populations, joint first authors Hyeseung Lee and Robert Fenster isolated tagged ribosomes that were expressed only in a given cell type, and sequenced the messenger RNA bound to them. In striatal neurons in the R6/2 mouse model of HD, at a time point when the mice were symptomatic, the researchers found more than 5,000 dysregulated genes. Curiously, some of the most upregulated transcripts were mitochondrial. Normally, mitochondrial RNA stays inside these organelles and is translated there. Finding large quantities of mitochondrial RNA stuck to cytoplasmic ribosomes indicates a breakdown in the mitochondria’s integrity or function.

How early does this start? The researchers turned to a series of knock-in HD mouse models that express huntingtin with CAG repeats of varying lengths. At a presymptomatic time point, mitochondrial RNA was already present in the cytoplasm of spiny projection neurons, in amounts correlating with the length of the CAG expansion.

Inflammatory RNA. Neurons (red) in control mice (top) do not express the innate immune sensor PKR (green), but the gene turns on in the presence of mutant huntingtin (bottom). Nuclei are blue. [Courtesy of Lee et al., Neuron.]

Previous research has found that, when in the cytoplasm, mitochondrial RNA can harm cells by triggering an immune response to foreign RNA that evolved to protect cells from viruses (Dhir et al., 2018). Foreign RNA binds and activates protein kinase R; in turn, PKR activates the interferon response, switching on proinflammatory cytokine production (Kim et al., 2018). PKR also shuts down translation and can spark apoptosis, both of which would also protect the brain from an invading virus.

This inflammatory response appears to get turned on in spiny projection neurons of the knock-in HD mice. The mitochondrial RNA immunoprecipitated with PKR, i.e., directly bound it, and interferon response genes were elevated.

What allowed the mitochondrial RNA to get out in the first place? Possibly it was sluggish mitochondrial energy production. Few genes were altered this early on—for example, only 119 in spiny projection neurons of a Q170-repeat mouse—but chief among those was dampened expression of genes fueling oxidative phosphorylation. In cultured neurons, putting pharmacologic brakes on oxidative phosphorylation induced PKR, the authors report. Neurons normally do not express PKR (see image above). The study’s genetic data implied that the transcriptional regulator Rarb might possibly constitute a druggable target; its activity would have to be activated.

Altogether, the data indicate that early deficits in oxidative phosphorylation in spiny projection neurons may lead to spillage of RNA from these energy-producing organelles, which ignites an inflammatory response.—Madolyn Bowman Rogers and Gabrielle Strobel

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References

Paper Citations

  1. Dhir A, Dhir S, Borowski LS, Jimenez L, Teitell M, Rötig A, Crow YJ, Rice GI, Duffy D, Tamby C, Nojima T, Munnich A, Schiff M, de Almeida CR, Rehwinkel J, Dziembowski A, Szczesny RJ, Proudfoot NJ. Mitochondrial double-stranded RNA triggers antiviral signalling in humans. Nature. 2018 Aug;560(7717):238-242. Epub 2018 Jul 25 PubMed.
  2. Kim Y, Park J, Kim S, Kim M, Kang MG, Kwak C, Kang M, Kim B, Rhee HW, Kim VN. PKR Senses Nuclear and Mitochondrial Signals by Interacting with Endogenous Double-Stranded RNAs. Mol Cell. 2018 Sep 20;71(6):1051-1063.e6. Epub 2018 Aug 30 PubMed.

Further Reading

Primary Papers

  1. Lee H, Fenster RJ, Pineda SS, Gibbs WS, Mohammadi S, Davila-Velderrain J, Garcia FJ, Therrien M, Novis HS, Gao F, Wilkinson H, Vogt T, Kellis M, LaVoie MJ, Heiman M. Cell Type-Specific Transcriptomics Reveals that Mutant Huntingtin Leads to Mitochondrial RNA Release and Neuronal Innate Immune Activation. Neuron. 2020 Sep 9;107(5):891-908.e8. Epub 2020 Jul 17 PubMed.

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