A glycolytic metabolite restores DNA repair activity of polynucleotide kinase 3-phosphatase in polyglutamine (PolyQ) diseases

Abstract

Huntingtons disease (HD) and spinocerebellar ataxia type 3 (SCA3) are the two most prevalent polyglutamine (polyQ) neurodegenerative diseases, caused by CAG (encoding glutamine) repeat expansion in the coding region of the huntingtin (HTT) and ataxin-3 (ATXN3) proteins, respectively. We have earlier reported that the activity, but not the protein level, of an essential DNA repair enzyme, polynucleotide kinase 3-phosphatase (PNKP), is severely abrogated in both HD and SCA3 resulting in accumulation of double-strand breaks in patients brain genome. While investigating the mechanistic basis for the loss of PNKP activity and accumulation of DNA double-strand breaks leading to neuronal death, we observed that PNKP interacts with the nuclear isoform of 6-phosphofructo-2-kinase fructose-2,6-bisphosphatase 3 (PFKFB3). Depletion of PFKFB3 markedly abrogates PNKP activity without changing its protein level. Notably, the levels of both PFKFB3 and its product fructose-2,6 bisphosphate (F2,6BP), an allosteric modulator of glycolysis, are significantly lower in the nuclear extracts of post-mortem brain tissues of HD and SCA3 patients. Supplementation of F2,6BP restored PNKP activity in the nuclear extracts of patients brain. Moreover, intracellular delivery of F2,6BP restored both the activity of PNKP and the integrity of transcribed genome in neuronal cells derived from striatum of HD mouse. Importantly, supplementing F2,6BP rescued the HD phenotype in Drosophila, suggesting F2,6BP to serve in vivo as a cofactor for the proper functionality of PNKP and thereby, of brain health. Our results thus provide a compelling rationale for exploring the therapeutic use of F2,6BP and structurally related compounds for treating polyQ diseases. SignificanceTo unravel the biological basis for the loss of PNKP activity in HD and SCA3, the two most prevalent polyglutamine neurodegenerative disorders, we analyzed PNKP interactome and found that the nuclear isoform of a glycolytic enzyme PFKFB3 associated with PNKP and other repair proteins forming a multiprotein complex. Surprisingly, we found that PFKFB3 and its biosynthetic product, F2,6BP are significantly low in the affected region of patients brain. Exogenous addition of F2,6BP restored PNKP activity in patients brain nuclear extract. Moreover, supplementing F2,6BP in HD cells and fruit flies restored genome integrity and rescued the disease symptoms. While there is no curative therapy for HD/SCA3, except symptom management, our discovery suggests that F2,6BP supplementation would be a promising therapeutic option.