{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,13]],"date-time":"2026-04-13T11:38:29Z","timestamp":1776080309482,"version":"3.50.1"},"reference-count":54,"publisher":"Oxford University Press (OUP)","issue":"12","license":[{"start":{"date-parts":[[2023,8,31]],"date-time":"2023-08-31T00:00:00Z","timestamp":1693440000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/pages\/standard-publication-reuse-rights"}],"funder":[{"DOI":"10.13039\/501100001691","name":"JSPS","doi-asserted-by":"publisher","award":["JP19K07343"],"award-info":[{"award-number":["JP19K07343"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"JSPS","doi-asserted-by":"publisher","award":["JP22K06878 (AF)"],"award-info":[{"award-number":["JP22K06878 (AF)"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"JSPS","doi-asserted-by":"publisher","award":["JP19H03203 (KN)"],"award-info":[{"award-number":["JP19H03203 (KN)"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"JSPS","doi-asserted-by":"publisher","award":["JP17H04036"],"award-info":[{"award-number":["JP17H04036"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"JSPS","doi-asserted-by":"publisher","award":["JP21H02678"],"award-info":[{"award-number":["JP21H02678"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100009619","name":"AMED","doi-asserted-by":"publisher","award":["JP22ym0126803 (AF)"],"award-info":[{"award-number":["JP22ym0126803 (AF)"]}],"id":[{"id":"10.13039\/100009619","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2023,12,14]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:p>Articular cartilage plays vital roles as a friction minimizer and shock absorber during joint movement but has a poor capacity to self-repair when damaged through trauma or disease. Cartilage tissue engineering is an innovative technique for cartilage regeneration, yet its therapeutic application requires chondrocytes in large numbers. Direct reprogramming of somatic cells to chondrocytes by expressing SOX9, KLF4, and c-MYC offers a promising option to generate chondrocytes in sufficient numbers; however, the low efficiency of the reprogramming system warrants further improvement. Here we referred to structural and functional features of SOX9 and performed alanine-scanning mutagenesis of functionally critical residues in the HMG box and at putative posttranslational modification (PTM) sites. We discovered that a SOX9 variant H131A\/K398A, doubly mutated in the HMG box (H131) and at a PTM site (K398), significantly upregulated expression of chondrogenic genes and potently induced chondrocytes from mouse embryonic fibroblasts. The H131A\/K398A variant remained unsumoylated in cells and exhibited a stronger DNA-binding activity than wild-type SOX9, especially when complexed with other proteins. Our results show that the novel SOX9 variant may be useful for efficient induction of chondrocytes and illuminate the strategic feasibility of mutating a transcription factor at functionally critical residues to expedite discovery of an optimized reprogramming factor.<\/jats:p>","DOI":"10.1093\/stmcls\/sxad066","type":"journal-article","created":{"date-parts":[[2023,8,31]],"date-time":"2023-08-31T15:42:22Z","timestamp":1693496542000},"page":"1157-1170","source":"Crossref","is-referenced-by-count":3,"title":["Engineering Critical Residues of SOX9 Discovers a Variant With Potent Capacity to Induce Chondrocytes"],"prefix":"10.1093","volume":"41","author":[{"given":"Yuya","family":"Sekiguchi","sequence":"first","affiliation":[{"name":"Laboratory of Gene Regulation, Institute of Medicine, University of Tsukuba , Tsukuba, Ibaraki , Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4960-7028","authenticated-orcid":false,"given":"Aya","family":"Fukuda","sequence":"additional","affiliation":[{"name":"Laboratory of Gene Regulation, Institute of Medicine, University of Tsukuba , Tsukuba, Ibaraki , Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0716-4142","authenticated-orcid":false,"given":"Ken","family":"Nishimura","sequence":"additional","affiliation":[{"name":"Laboratory of Gene Regulation, Institute of Medicine, University of Tsukuba , Tsukuba, Ibaraki , Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Koji","family":"Hisatake","sequence":"additional","affiliation":[{"name":"Laboratory of Gene Regulation, Institute of Medicine, University of Tsukuba , Tsukuba, Ibaraki , Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"286","published-online":{"date-parts":[[2023,8,31]]},"reference":[{"key":"2023121512033456000_CIT0001","first-page":"461","article-title":"The basic science of articular cartilage: structure, composition, and function","volume-title":"Sports Health","author":"Sophia Fox","year":"2009"},{"key":"2023121512033456000_CIT0002","first-page":"77","article-title":"Articular cartilage: structure, injuries and review of management","volume-title":"Br Med Bull","author":"Bhosale","year":"2008"},{"key":"2023121512033456000_CIT0003","first-page":"46","article-title":"Comparative effectiveness of pharmacologic interventions for knee osteoarthritis: a systematic review and network meta-analysis","volume-title":"Ann Intern Med","author":"Bannuru","year":"2015"},{"key":"2023121512033456000_CIT0004","first-page":"889","article-title":"Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation","volume-title":"New Eng J Med","author":"Brittberg","year":"1994"},{"issue":"1","key":"2023121512033456000_CIT0005","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1177\/03635465020300011601","article-title":"Autologous chondrocyte transplantation: biomechanics and long-term durability","volume":"30","author":"Peterson","year":"2002","journal-title":"Am J Sports Med"},{"key":"2023121512033456000_CIT0006","first-page":"623","article-title":"Articular cartilage aging-potential regenerative capacities of cell manipulation and stem cell therapy","volume-title":"Int J Mol Sci","author":"Krajewska-W\u0142odarczyk","year":"2018"},{"key":"2023121512033456000_CIT0007","first-page":"476","article-title":"Age related changes in human articular chondrocyte yield, proliferation and post-expansion chondrogenic capacity","volume-title":"Osteoarthr and Cartil","author":"Barbero","year":"2004"},{"key":"2023121512033456000_CIT0008","first-page":"640","article-title":"Generation of hyaline cartilaginous tissue from mouse adult dermal fibroblast culture by defined factors","volume-title":"J Clin Invest","author":"Hiramatsu","year":"2011"},{"key":"2023121512033456000_CIT0009","first-page":"3","article-title":"Signaling pathways regulating cartilage growth plate formation and activity","volume-title":"Semin Cell Dev Biol","author":"Samsa","year":"2017"},{"issue":"1","key":"2023121512033456000_CIT0010","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1038\/ng0195-15","article-title":"The Sry-related gene Sox9 is expressed during chondrogenesis in mouse embryos","volume":"9","author":"Wright\u2019","year":"1995","journal-title":"Nat Genet"},{"key":"2023121512033456000_CIT0011","first-page":"1111","article-title":"Autosomal sex reversal and campomelic dysplasia are caused by mutations q and around the SRY-related gene 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chondrocyte-specific enhancer","volume-title":"Mol Cell Biol","author":"Huang","year":"2000"},{"key":"2023121512033456000_CIT0024","first-page":"499","article-title":"The transcription factor Sox9 is degraded by the ubiquitin-proteasome system and stabilized by a mutation in a ubiquitin-target site","volume-title":"Matrix Biol","author":"Akiyama","year":"2005"},{"key":"2023121512033456000_CIT0025","first-page":"191","article-title":"Rho kinase\u2013dependent activation of SOX9 in chondrocytes","volume-title":"Arthritis Rheum","author":"Haudenschild","year":"2010"},{"key":"2023121512033456000_CIT0026","first-page":"2882","article-title":"Phosphorylation of Sox9 is required for neural crest delamination and is regulated downstream of BMP and canonical Wnt signaling","volume-title":"Proc Natl Acad Sci USA","author":"Liu","year":"2013"},{"key":"2023121512033456000_CIT0027","first-page":"3011","article-title":"Transcriptional regulation of chondrogenesis by coactivator Tip60 via chromatin association with Sox9 and Sox5","volume-title":"Nucleic Acids Res","author":"Hattori","year":"2008"},{"key":"2023121512033456000_CIT0028","first-page":"4034","article-title":"Chondrocyte hypertrophy in osteoarthritis: mechanistic studies and models for the identification of new therapeutic strategies","volume-title":"Cells","author":"Chawla","year":"2022"},{"key":"2023121512033456000_CIT0029","first-page":"216","article-title":"Hypertrophic differentiation of chondrocytes in osteoarthritis: the developmental aspect of degenerative joint disorders","volume-title":"Arthritis Res Ther","author":"Dreier","year":"2010"},{"key":"2023121512033456000_CIT0030","first-page":"8183","article-title":"The transcription factors SOX9 and SOX5\/SOX6 cooperate genome-wide through super-enhancers to drive chondrogenesis","volume-title":"Nucleic Acids Res","author":"Liu","year":"2015"},{"key":"2023121512033456000_CIT0031","first-page":"S69","article-title":"L-Sox5, Sox6 and Sox9 control essential steps of the chondrocyte differentiation pathway","volume-title":"Osteoarthr and Cartil","author":"Lefebvre","year":"2001"},{"key":"2023121512033456000_CIT0032","first-page":"122","article-title":"Cancer-associated missense mutations enhance the pluripotency reprogramming activity of OCT4 and SOX17","volume-title":"FEBS J","author":"Srivastava","year":"2020"},{"key":"2023121512033456000_CIT0033","first-page":"593","article-title":"Directed evolution of reprogramming factors by cell selection and sequencing","volume-title":"Stem Cell Rep","author":"Veerapandian","year":"2018"},{"key":"2023121512033456000_CIT0034","first-page":"103525","article-title":"Structurally-discovered KLF4 variants accelerate and stabilize reprogramming to pluripotency","volume-title":"iScience","author":"Borisova","year":"2022"},{"key":"2023121512033456000_CIT0035","first-page":"4666","article-title":"Structure-based discovery of NANOG variant with enhanced properties to promote self-renewal and reprogramming of pluripotent stem cells","volume-title":"Proc Natl Acad Sci U S A","author":"Hayashi","year":"2015"},{"key":"2023121512033456000_CIT0036","first-page":"3922","article-title":"Structure and decoy-mediated inhibition of the SOX18\/ Prox1 -DNA interaction","volume-title":"Nucleic Acids Res.","author":"Klaus","year":"2016"},{"key":"2023121512033456000_CIT0037","first-page":"705","article-title":"Molecular basis of human 46X,Y sex reversal revealed from the three-dimensional solution structure of the human SRY-DNA complex","volume-title":"Cell","author":"Werner","year":"1995"},{"key":"2023121512033456000_CIT0038","first-page":"619","article-title":"The structure of Sox17 bound to DNA reveals a conserved bending topology but selective protein interaction platforms","volume-title":"J Mol Biol","author":"Palasingam","year":"2009"},{"key":"2023121512033456000_CIT0039","first-page":"39","article-title":"The crystal structure of the Sox4 HMG domain\u2013DNA complex suggests a mechanism for positional interdependence in DNA recognition","volume-title":"Biochem J","author":"Jauch","year":"2012"},{"key":"2023121512033456000_CIT0040","first-page":"e0161432","article-title":"A model for dimerization of the SOX Group E transcription factor family","volume-title":"PLoS One","author":"Ramsook","year":"2016"},{"key":"2023121512033456000_CIT0041","first-page":"2048","article-title":"Crystal structure of a POU\/HMG\/DNA ternary complex suggests differential assembly of Oct4 and Sox2 on two enhancers","volume-title":"Genes Dev","author":"Rem\u00e9nyi","year":"2003"},{"key":"2023121512033456000_CIT0042","first-page":"938","article-title":"Turning a hobby into a job: How duplicated genes find new functions","volume-title":"Nat Rev Genet","author":"Conant","year":"2008"},{"key":"2023121512033456000_CIT0043","first-page":"97","article-title":"The evolution of gene duplications: classifying and distinguishing between models","volume-title":"Nat Rev Genet","author":"Innan","year":"2010"},{"key":"2023121512033456000_CIT0044","first-page":"112","article-title":"High-throughput affinity measurements of transcription factor and DNA mutations reveal affinity and specificity determinants","volume-title":"Cell Syst","author":"Aditham","year":"2021"},{"key":"2023121512033456000_CIT0045","first-page":"1372","article-title":"Selective deployment of transcription factor paralogs with submaximal strength facilitates gene regulation in the immune system","volume-title":"Nat Immunol","author":"Bruno","year":"2019"},{"key":"2023121512033456000_CIT0046","first-page":"12654","article-title":"SUMO-1 conjugation in vivo requires both a consensus modification motif and nuclear targeting","volume-title":"J Biol Chem","author":"Rodriguez","year":"2001"},{"key":"2023121512033456000_CIT0047","first-page":"593","article-title":"SoxE factors function equivalently during neural crest and inner ear development and their activity is regulated by SUMOylation","volume-title":"Dev Cell","author":"Taylor","year":"2005"},{"key":"2023121512033456000_CIT0048","first-page":"799","article-title":"SUMOylated SoxE factors recruit Grg4 and function as transcriptional repressors in the neural crest","volume-title":"J Cell Biol","author":"Lee","year":"2012"},{"key":"2023121512033456000_CIT0049","first-page":"27224","article-title":"Transcriptional co-activators CREB-binding protein and p300 regulate chondrocyte-specific gene expression via association with Sox9","volume-title":"J Biol Chem","author":"Tsuda","year":"2003"},{"key":"2023121512033456000_CIT0050","first-page":"1307","article-title":"A conserved role for transcription factor sumoylation in binding-site selection","volume-title":"Curr Genet","author":"Rosonina","year":"2019"},{"key":"2023121512033456000_CIT0051","first-page":"1575","article-title":"SUMOylation regulates the chromatin occupancy and anti-proliferative gene programs of glucocorticoid receptor","volume-title":"Nucleic Acids Res","author":"Paakinaho","year":"2014"},{"key":"2023121512033456000_CIT0052","first-page":"8310","article-title":"SUMOylation modulates the transcriptional activity of androgen receptor in a target gene and pathway selective manner","volume-title":"Nucleic Acids Res","author":"Sutinen","year":"2014"},{"key":"2023121512033456000_CIT0053","first-page":"2679","article-title":"Pioneer transcription factors in cell reprogramming","volume-title":"Genes Dev","author":"Iwafuchi-Doi","year":"2014"},{"key":"2023121512033456000_CIT0054","first-page":"704","article-title":"A serial shRNA screen for roadblocks to reprogramming identifies the protein modifier SUMO2","volume-title":"Stem Cell Rep","author":"Borkent","year":"2016"}],"container-title":["Stem 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