Functional Characterisation of the Circular RNA, circHTT(2-6), in Huntington's Disease

Laura Gantley; Brett W Stringer; Vanessa M Conn; Youichirou Ootsuka; Duncan Holds; Mark Slee; Kamelya Aliakbari; Kirsty Kirk; Rebecca J Ormsby; Stuart T Webb; Adrienne Hanson; He Lin; Luke A Selth; Simon J Conn

doi:10.3390/cells12091337

Functional Characterisation of the Circular RNA, circHTT(2-6), in Huntington's Disease

Cells. 2023 May 7;12(9):1337. doi: 10.3390/cells12091337.

Authors

Laura Gantley¹, Brett W Stringer^{1

2}, Vanessa M Conn¹, Youichirou Ootsuka^{1

3}, Duncan Holds⁴, Mark Slee¹, Kamelya Aliakbari¹, Kirsty Kirk¹, Rebecca J Ormsby¹, Stuart T Webb¹, Adrienne Hanson¹, He Lin¹, Luke A Selth^{1

5

6}, Simon J Conn¹

Affiliations

¹ Flinders Health and Medical Research Institute (FHMRI), College of Medicine and Public Health, Flinders University, Bedford Park, SA 5042, Australia.
² Lifelong Health, South Australian Health and Medical Research Institute, Adelaide, SA 5000, Australia.
³ Centre for Neuroscience, Human Physiology, College of Medicine and Public Health, Flinders University, Bedford Park, SA 5042, Australia.
⁴ Department of Genetics and Molecular Pathology, SA Pathology, Flinders Medical Centre, Bedford Park, SA 5042, Australia.
⁵ Freemasons Centre for Male Health and Wellbeing, Flinders University, Bedford Park, SA 5042, Australia.
⁶ Adelaide Medical School, University of Adelaide, Adelaide, SA 5000, Australia.

Abstract

Trinucleotide repeat disorders comprise ~20 severe, inherited, human neuromuscular and neurodegenerative disorders, which result from an abnormal expansion of repetitive sequences in the DNA. The most common of these, Huntington's disease (HD), results from expansion of the CAG repeat region in exon 1 of the HTT gene via an unknown mechanism. Since non-coding RNAs have been implicated in the initiation and progression of many diseases, herein we focused on a circular RNA (circRNA) molecule arising from non-canonical splicing (backsplicing) of HTT pre-mRNA. The most abundant circRNA from HTT, circHTT(2-6), was found to be more highly expressed in the frontal cortex of HD patients, compared with healthy controls, and positively correlated with CAG repeat tract length. Furthermore, the mouse orthologue (mmu_circHTT(2-6)) was found to be enriched within the brain and specifically the striatum, a region enriched for medium spiny neurons that are preferentially lost in HD. Transgenic overexpression of circHTT(2-6) in two human cell lines-SH-SY5Y and HEK293-reduced cell proliferation and nuclear size without affecting cell cycle progression or cellular size, or altering the CAG repeat region length within HTT. CircHTT(2-6) overexpression did not alter total HTT protein levels, but reduced its nuclear localisation. As these phenotypic and genotypic changes resemble those observed in HD patients, our results suggest that circHTT(2-6) may play a functional role in the pathophysiology of this disease.

Keywords: HTT; Huntingtin; Huntington’s disease; circular RNA; triplet repeat disorders.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Animals, Genetically Modified
HEK293 Cells
Humans
Huntington Disease* / metabolism
Mice
Neuroblastoma*
RNA, Circular / genetics

Substances

RNA, Circular

Grants and funding

This research was funded by the National Health and Medical Research Council, GNT1190814, the Australian Research Council Future Fellowship Scheme, FT160100318 and the Tour de Cure grant, Tour de Cure: RSP-089-202, to S.J.C.; the Flinders Foundation Health Seed Grant to S.J.C., V.M.C., and B.W.S. L.A.S. was supported by a Principal Cancer Research Fellowship (PRF2919) awarded by the Cancer Council’s Beat Cancer project on behalf of its donors, the State Government through the Department of Health, and the Australian Government through the Medical Research Future Fund.