{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T05:57:09Z","timestamp":1782280629327,"version":"3.54.5"},"posted":{"date-parts":[[2024,11,4]]},"group-title":"Neuroscience","reference-count":39,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,11,4]],"date-time":"2024-11-04T00:00:00Z","timestamp":1730678400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nd\/4.0\/"}],"funder":[{"name":"NIH","award":["R56NS073976"],"award-info":[{"award-number":["R56NS073976"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,6,30]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Several reports have indicated that impaired mitochondrial function contributes to the development and progression of Huntington\u2019s disease (HD). Mitochondrial genome damage, particularly DNA strand breaks, is a potential cause for its compromised functionality. Here we show that the activity of polynucleotide kinase 3\u2019-phosphatase (PNKP), a critical DNA end-processing enzyme, is significantly decreased in the mitochondrial extract of HD patients\u2019 brains due to a lower level of fructose-2,6 bisphosphate (F2,6BP), a biosynthetic product of 6-phosphofructo-2-kinase fructose-2,6-bisphosphatase 3 (PFKFB3). Such decrease in PNKP activity leads to persistent DNA strand breaks that are refractory to subsequent steps for repair completion. Both PFKFB3 and F2,6BP, an allosteric modulator of glycolysis, are also present in the mitochondria and PFKFB3 is part of a mitochondrial DNA repair complex containing HTT, PNKP, DNA Pol \u03b3 (POLG) and Lig III\u03b1. Notably, PNKP binds F2,6BP (Kd= 525\u00b125 nM) and utilizes it as a cofactor. The levels of both F2,6BP and PFKFB3 are significantly decreased in the mitochondrial extract of HD mouse striatal neuronal cells and patients\u2019 brain. Activity of PNKP is thus severely decreased in the mitochondrial extract; however, addition of F2,6BP restored its activity. Moreover, supplementation of F2,6BP in HD cells restored PFKFB3 level, mitochondrial genome integrity and partially restored mitochondrial membrane potential, mitochondrial respiration and prevented pathogenic aggregate formation. We also observed that supplementation with F2,6BP restored mitochondrial genome integrity in an HD\n                  <jats:italic>Drosophila<\/jats:italic>\n                  model. Our findings, therefore, suggest that F2,6BP-mediated restoration of PNKP activity could have a profound impact in ameliorating neurodegenerative symptoms in HD.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Significance<\/jats:title>\n                  <jats:p>We reported earlier the loss of PNKP activity in the nuclear extracts from HD patients\u2019 brain. However, a glycolytic metabolite, F2,6BP, can restore PNKP activity and rescue organismal phenotypes in HD fly models. As PNKP is present in mitochondria and several reports indicate that mitochondrial dysfunction contributes to HD, we therefore analyzed PNKP activity in the mitochondrial extract. Surprisingly, we found that PFKFB3 and its product, F2,6BP are present in mitochondria, but significantly low in patients\u2019 brains. Exogenous addition of F2,6BP restored PNKP activity in patients\u2019 brain mitochondrial extract. Moreover, supplementing F2,6BP in HD cells and fruit flies restored mitochondrial genome integrity suggesting maintaining adequate intracellular F2,6BP levels is critical for proper functionality of PNKP and thereby of brain health.<\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2024.11.04.621834","type":"posted-content","created":{"date-parts":[[2024,11,4]],"date-time":"2024-11-04T21:05:18Z","timestamp":1730754318000},"source":"Crossref","is-referenced-by-count":2,"title":["F2,6BP restores mitochondrial genome integrity in Huntington\u2019s Disease"],"prefix":"10.64898","author":[{"given":"Anirban","family":"Chakraborty","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Santi 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