{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T18:59:24Z","timestamp":1785437964201,"version":"3.56.0"},"posted":{"date-parts":[[2026,3,17]]},"group-title":"Neuroscience","reference-count":37,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,3,17]],"date-time":"2026-03-17T00:00:00Z","timestamp":1773705600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000065","name":"National Institute of Neurological Disorders and Stroke","doi-asserted-by":"crossref","award":["1R35NS127248-01"],"award-info":[{"award-number":["1R35NS127248-01"]}],"id":[{"id":"10.13039\/100000065","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000065","name":"National Institute of Neurological Disorders and Stroke","doi-asserted-by":"crossref","award":["R61NS11075"],"award-info":[{"award-number":["R61NS11075"]}],"id":[{"id":"10.13039\/100000065","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000065","name":"National Institute of Neurological Disorders and Stroke","doi-asserted-by":"crossref","award":["R33NS11075"],"award-info":[{"award-number":["R33NS11075"]}],"id":[{"id":"10.13039\/100000065","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000065","name":"National Institute of Neurological Disorders and Stroke","doi-asserted-by":"crossref","award":["2R15NS116511-02"],"award-info":[{"award-number":["2R15NS116511-02"]}],"id":[{"id":"10.13039\/100000065","id-type":"DOI","asserted-by":"crossref"}]},{"award":["GM044757"],"award-info":[{"award-number":["GM044757"]}],"id":[{"id":"https:\/\/ror.org\/04q48ey07","id-type":"ROR","asserted-by":"publisher"}]},{"award":["Early Career Award"],"award-info":[{"award-number":["Early Career Award"]}],"id":[{"id":"https:\/\/ror.org\/03s9xxw55","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,3,17]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  The spinocerebellar ataxia type 5 (SCA5) L253P mutation in \u03b2-III-spectrin causes high-affinity actin binding. Here we developed a CRISPR knock-in mouse to determine the i\n                  <jats:italic>n vivo<\/jats:italic>\n                  impact of L253P on Purkinje neurons and motor activity, and to establish a model for future testing of SCA5 therapeutics. Significantly, the knock-in mouse shows impaired motor activity on elevated beam assays at 20 weeks. In the cerebellum, L253P causes a subcellular redistribution of \u03b2-III-spectrin in Purkinje neurons. This is marked by loss of \u03b2-III-spectrin in distal dendrites, accumulation of \u03b2-III-spectrin at the plasma membrane of the soma and proximal dendrites, and formation of inclusions in the soma. The inclusions additionally contain F-actin and \u03b1-II-spectrin, accumulate around the nucleus, form at an early age, and are larger in homozygous\n                  <jats:italic>\u03b2-III-spectrin<\/jats:italic>\n                  <jats:sup>\n                    <jats:italic>L253P\/L253P<\/jats:italic>\n                  <\/jats:sup>\n                  compared to heterozygous\n                  <jats:italic>\u03b2-III-spectrin<\/jats:italic>\n                  <jats:sup>\n                    <jats:italic>L253P\/+<\/jats:italic>\n                  <\/jats:sup>\n                  mice. In contrast, neurons of the hippocampus and cerebral cortex, where \u03b2-III-spectrin is also known to be expressed, abnormally accumulate \u03b2-III-spectrin at the plasma membrane but do not form inclusions. To gain greater insight into disease mechanisms, unbiased proteomics identified over 150 cerebellar proteins that physically associate with \u03b2-III-spectrin. Of these, cluster analysis revealed a group of 41 proteins, including glutamate receptors, SERCA2, and CaMKII, linked to synaptic transmission. Thus, the effect of the L253P to alter \u03b2-III-spectrin localization, including decreased levels in distal dendrites, is likely associated with a disruption of \u03b2-III-spectrin function in postsynaptic signaling. Consistent with this, and in agreement with prior findings in knockout mice, the L253P\n                  <jats:italic>\u03b2-III-spectrin<\/jats:italic>\n                  knock-in mouse here shows that CaMKII, a calcium sensor and key mediator of glutamate signaling, is ~2-fold activated. Further, the abundance of EAAT4, a glutamate transporter, is significantly reduced. The L253P knock-in mouse primes future preclinical testing of SCA5 therapeutics, such as small molecule modulators of spectrin-actin binding, and glutamate and calcium signaling pathways.\n                <\/jats:p>","DOI":"10.64898\/2026.03.14.711824","type":"posted-content","created":{"date-parts":[[2026,3,17]],"date-time":"2026-03-17T09:35:12Z","timestamp":1773740112000},"source":"Crossref","is-referenced-by-count":0,"title":["Impaired motor activity in a CRISPR SCA5 L253P knock-in mouse is associated with selective \u03b2-III-spectrin subcellular redistribution in the cerebellum"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-8085-1822","authenticated-orcid":false,"given":"Adam W.","family":"Avery","sequence":"first","affiliation":[{"name":"Department of Chemistry, Oakland University, Rochester, MI 48309, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0800-5856","authenticated-orcid":false,"given":"Brennon L.","family":"O\u2019Callaghan","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, Institute for Translational Neuroscience, University of Minnesota, Minneapolis, MN 55455, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthew T.","family":"Thiel","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Oakland University, Rochester, MI 48309, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5341-5512","authenticated-orcid":false,"given":"Sarah A.","family":"Denha","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Oakland University, Rochester, MI 48309, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-2905-7726","authenticated-orcid":false,"given":"Devon G.","family":"O\u2019Callaghan","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, Institute for Translational Neuroscience, University of Minnesota, Minneapolis, MN 55455, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-5070-6881","authenticated-orcid":false,"given":"Emma M.","family":"Cismas","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Oakland University, Rochester, MI 48309, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7187-7657","authenticated-orcid":false,"given":"Jared","family":"Lamp","sequence":"additional","affiliation":[{"name":"Integrated Mass Spectrometry Unit, College of Human Medicine, Michigan State University, Grand Rapids, MI 49503, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6118-741X","authenticated-orcid":false,"given":"Harry T.","family":"Orr","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, Institute for Translational Neuroscience, University of Minnesota, Minneapolis, MN 55455, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4911-6042","authenticated-orcid":false,"given":"Thomas S.","family":"Hays","sequence":"additional","affiliation":[{"name":"Department of Genetics and Cell Biology, University of Minnesota, Minneapolis, MN 55455, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026031912551197000_2026.03.14.711824v1.1","doi-asserted-by":"publisher","DOI":"10.1038\/ng1728"},{"key":"2026031912551197000_2026.03.14.711824v1.2","doi-asserted-by":"publisher","DOI":"10.1212\/01.WNL.0000103293.63340.C1"},{"key":"2026031912551197000_2026.03.14.711824v1.3","doi-asserted-by":"publisher","DOI":"10.1016\/S0169-328X(98)00068-0"},{"key":"2026031912551197000_2026.03.14.711824v1.4","doi-asserted-by":"crossref","first-page":"21375","DOI":"10.1038\/srep21375","article-title":"A human beta-III-spectrin spinocerebellar ataxia type 5 mutation causes high-affinity F-actin binding","volume":"6","year":"2016","journal-title":"Scientific reports"},{"key":"2026031912551197000_2026.03.14.711824v1.5","doi-asserted-by":"crossref","first-page":"1350","DOI":"10.1038\/s41467-017-01367-w","article-title":"Structural basis for high-affinity actin binding revealed by a beta-III-spectrin SCA5 missense mutation","volume":"8","year":"2017","journal-title":"Nature communications"},{"key":"2026031912551197000_2026.03.14.711824v1.6","doi-asserted-by":"crossref","unstructured":"Atang, A. E. , Keller, A. R. , Denha, S. A. , and Avery, A. W. (2023) Increased Actin Binding Is a Shared Molecular Consequence of Numerous SCA5 Mutations in beta-III-Spectrin Cells 12,","DOI":"10.3390\/cells12162100"},{"key":"2026031912551197000_2026.03.14.711824v1.7","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.RA120.015417"},{"key":"2026031912551197000_2026.03.14.711824v1.8","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2023.102956102956"},{"key":"2026031912551197000_2026.03.14.711824v1.9","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1705043114"},{"key":"2026031912551197000_2026.03.14.711824v1.10","doi-asserted-by":"crossref","unstructured":"Fujishima, K. , Kurisu, J. , Yamada, M. , and Kengaku, M. (2020) betaIII spectrin controls the planarity of Purkinje cell dendrites by modulating perpendicular axon-dendrite interactions Development 147,","DOI":"10.1242\/dev.194530"},{"key":"2026031912551197000_2026.03.14.711824v1.11","doi-asserted-by":"crossref","first-page":"110350","DOI":"10.1016\/j.jbc.2025.110350","article-title":"Molecular consequences of SCA5 mutations in the spectrin-repeat domains of beta-III-spectrin","volume":"301","year":"2025","journal-title":"J Biol Chem"},{"key":"2026031912551197000_2026.03.14.711824v1.12","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1001522107"},{"key":"2026031912551197000_2026.03.14.711824v1.13","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.3520-16.2017"},{"key":"2026031912551197000_2026.03.14.711824v1.14","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1707108114"},{"key":"2026031912551197000_2026.03.14.711824v1.15","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.6065-09.2010"},{"key":"2026031912551197000_2026.03.14.711824v1.16","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.3332-11.2011"},{"key":"2026031912551197000_2026.03.14.711824v1.17","first-page":"4448","article-title":"Posterior cerebellar Purkinje cells in an SCA5\/SPARCA1 mouse model are especially vulnerable to the synergistic effect of loss of beta-III spectrin and GLAST","volume":"25","year":"2016","journal-title":"Hum Mol Genet"},{"key":"2026031912551197000_2026.03.14.711824v1.18","doi-asserted-by":"publisher","DOI":"10.1093\/hmg\/ddy169"},{"key":"2026031912551197000_2026.03.14.711824v1.19","doi-asserted-by":"crossref","first-page":"20990","DOI":"10.1038\/s41598-025-05511-1","article-title":"Automated gait analysis indicates efficacy of T-type calcium channel inhibition for mitigation of disrupted calcium signalling in an SCA5 mouse model","volume":"15","year":"2025","journal-title":"Sci Rep"},{"key":"2026031912551197000_2026.03.14.711824v1.20","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.0876-14.2014"},{"key":"2026031912551197000_2026.03.14.711824v1.21","doi-asserted-by":"publisher","DOI":"10.1038\/35065091"},{"key":"2026031912551197000_2026.03.14.711824v1.22","doi-asserted-by":"publisher","DOI":"10.1093\/hmg\/ddu103"},{"key":"2026031912551197000_2026.03.14.711824v1.23","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.1132-21.2021"},{"key":"2026031912551197000_2026.03.14.711824v1.24","doi-asserted-by":"publisher","DOI":"10.1093\/NAR\/GKAC1000"},{"key":"2026031912551197000_2026.03.14.711824v1.25","doi-asserted-by":"crossref","first-page":"866","DOI":"10.1016\/j.ajhg.2017.10.008","article-title":"Dominant Mutations in GRM1 Cause Spinocerebellar Ataxia Type 44","volume":"101","year":"2017","journal-title":"Am J Hum Genet"},{"key":"2026031912551197000_2026.03.14.711824v1.26","doi-asserted-by":"publisher","DOI":"10.1001\/archneur.60.12.1749"},{"key":"2026031912551197000_2026.03.14.711824v1.27","doi-asserted-by":"publisher","DOI":"10.1038\/ng1194-280"},{"key":"2026031912551197000_2026.03.14.711824v1.28","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pgen.1003074"},{"key":"2026031912551197000_2026.03.14.711824v1.29","doi-asserted-by":"publisher","DOI":"10.1002\/ajmg.a.38332"},{"key":"2026031912551197000_2026.03.14.711824v1.30","doi-asserted-by":"publisher","DOI":"10.1038\/ejhg.2013.150"},{"key":"2026031912551197000_2026.03.14.711824v1.31","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.200905158"},{"key":"2026031912551197000_2026.03.14.711824v1.32","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05074-5"},{"key":"2026031912551197000_2026.03.14.711824v1.33","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-020-18825-7"},{"key":"2026031912551197000_2026.03.14.711824v1.34","doi-asserted-by":"publisher","DOI":"10.1126\/science.ado2032"},{"key":"2026031912551197000_2026.03.14.711824v1.35","doi-asserted-by":"publisher","DOI":"10.1126\/sciadv.1500301"},{"key":"2026031912551197000_2026.03.14.711824v1.36","doi-asserted-by":"publisher","DOI":"10.1172\/JCI43621"},{"key":"2026031912551197000_2026.03.14.711824v1.37","doi-asserted-by":"publisher","DOI":"10.1038\/s41583-022-00674-6"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.03.14.711824","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,19]],"date-time":"2026-03-19T19:55:21Z","timestamp":1773950121000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.03.14.711824"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,3,17]]},"references-count":37,"URL":"https:\/\/doi.org\/10.64898\/2026.03.14.711824","relation":{},"subject":[],"published":{"date-parts":[[2026,3,17]]},"subtype":"preprint"}}