{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,8,19]],"date-time":"2026-08-19T14:27:15Z","timestamp":1787149635134,"version":"build-2736575974"},"posted":{"date-parts":[[2025,9,4]]},"group-title":"Neuroscience","reference-count":63,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,9,4]],"date-time":"2025-09-04T00:00:00Z","timestamp":1756944000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"award":["P50HD104463, R01NS099280 and R01NS086810"],"award-info":[{"award-number":["P50HD104463, R01NS099280 and R01NS086810"]}],"id":[{"id":"https:\/\/ror.org\/01s5ya894","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,8,16]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Nucleotide repeat expansions support production of toxic peptides from putatively non-coding regions of the human genome without use of an AUG initiation codon. This process, known as RAN translation, was originally described at CAG repeats with products generated in all three possible reading frames. Recently, cryptic mis-splicing of mRNAs in which the CAG repeat acts as a splice acceptor was proposed as an alternative mechanism for RAN protein production. To investigate this, we generated DNA and RNA based reporters to assess RAN translation in the context of a CAG repeat expansion in exon 1 of the Huntingtin gene (\n                  <jats:italic>HTT),<\/jats:italic>\n                  associated with Huntington Disease. HTT CAG repeats support RAN translation in both the alanine (GCA) and glutamine (CAG) frames, with or without upstream AUG start codons or near-AUG cognate codons, and at levels comparable to RAN translation from CGG and GGGGCC repeats. CAG RAN translation products were readily detectable from\n                  <jats:italic>in vitro<\/jats:italic>\n                  transcribed RNAs transfected into human cells and rodent neurons - suggesting that plasmid based aberrant splicing into CAG repeats cannot fully explain the observed phenomena. These findings indicate that RAN translation from HTT CAG repeats shares key mechanistic parameters with other disease-associated repeat expansions.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Key Points<\/jats:title>\n                  <jats:list list-type=\"bullet\">\n                    <jats:list-item>\n                      <jats:p>Translation at HTT CAG repeats occurs in at least two reading frames absent any AUG start codon.<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Like RAN translation at other repeats, HTT CAG RAN translation is enhanced by cell stress.<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>HTT CAG RAN translation readily occurs from in vitro transcribed RNAs in neurons.<\/jats:p>\n                    <\/jats:list-item>\n                  <\/jats:list>\n                <\/jats:sec>","DOI":"10.1101\/2025.09.02.673827","type":"posted-content","created":{"date-parts":[[2025,9,5]],"date-time":"2025-09-05T07:20:14Z","timestamp":1757056814000},"source":"Crossref","is-referenced-by-count":0,"title":["Mechanistic insights into RAN translation within the Huntingtin coding sequence"],"prefix":"10.64898","author":[{"given":"Amy","family":"Krans","sequence":"first","affiliation":[{"name":"Department of Neurology, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard M.","family":"Albertson","sequence":"additional","affiliation":[{"name":"Department of Neurology, University of Michigan, Ann Arbor, MI 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Connor J.","family":"Maltby","sequence":"additional","affiliation":[{"name":"Department of Neurology, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Department of Neurology, University of Michigan, Ann Arbor, MI 48109, USA; now at Ulysses Neuroscience, Dublin, Republic of Ireland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4781-6376","authenticated-orcid":false,"given":"Peter K.","family":"Todd","sequence":"additional","affiliation":[{"name":"Department of Neurology, University of Michigan, Ann Arbor, MI 48109, USA"},{"name":"Ann Arbor Veterans Administration Healthcare, Ann Arbor, MI, 48109, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026081906501395000_2025.09.02.673827v2.1","doi-asserted-by":"publisher","DOI":"10.1111\/ene.13413"},{"key":"2026081906501395000_2025.09.02.673827v2.2","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(93)90585-E"},{"key":"2026081906501395000_2025.09.02.673827v2.3","doi-asserted-by":"publisher","DOI":"10.1038\/ng0893-398"},{"key":"2026081906501395000_2025.09.02.673827v2.4","doi-asserted-by":"publisher","DOI":"10.1038\/s41556-024-01414-x"},{"key":"2026081906501395000_2025.09.02.673827v2.5","doi-asserted-by":"publisher","DOI":"10.1126\/science.277.5334.1990"},{"key":"2026081906501395000_2025.09.02.673827v2.6","first-page":"910","article-title":"The Biology of Huntingtin","volume":"8","year":"2016","journal-title":"Neuron"},{"key":"2026081906501395000_2025.09.02.673827v2.7","doi-asserted-by":"publisher","DOI":"10.1038\/s41580-021-00382-6"},{"key":"2026081906501395000_2025.09.02.673827v2.8","doi-asserted-by":"publisher","DOI":"10.1186\/s13024-023-00642-3"},{"key":"2026081906501395000_2025.09.02.673827v2.9","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1013343108"},{"key":"2026081906501395000_2025.09.02.673827v2.10","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2013.02.004"},{"key":"2026081906501395000_2025.09.02.673827v2.11","first-page":"1335","article-title":"The C9orf72 GGGGCC repeat is translated into aggregating dipeptide-repeat proteins in FTLD\/ALS","volume":"33","year":"2013","journal-title":"Science"},{"key":"2026081906501395000_2025.09.02.673827v2.12","first-page":"1888","article-title":"Repeat-Associated Non-ATG (RAN) Translation in Fuchs\u2019 Endothelial Corneal Dystrophy","volume":"5","year":"2018","journal-title":"Invest Ophthalmol Vis Sci"},{"key":"2026081906501395000_2025.09.02.673827v2.13","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2013.03.026"},{"key":"2026081906501395000_2025.09.02.673827v2.14","doi-asserted-by":"crossref","unstructured":"Zu, T. , Cleary, J.D. , Liu, Y. , Banez-Coronel, M. , Bubenik, J.L. , Ayhan, F. , Ashizawa, T. , Xia, G. , Clark, H.B. , Yachnis, A.T. et al. 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