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Overexpression of chromobox homolog 4 (CBX4) in a mouse system was demonstrated to alleviate post-traumatic osteoarthritis (PTOA) by reducing cellular senescence. Additionally, replicative cellular senescence of WI-38 fibroblasts can be attenuated by CBX4. However, the mechanisms underlying this senomorphic function of CBX4 are not fully understood. In this study, we aimed to investigate the role of CBX4 in cellular senescence in human primary osteoarthritic chondrocytes and to identify the functional domains of CBX4 necessary for its function in modulating senescence.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Methods<\/jats:title>\n                <jats:p>Chondrocytes, isolated from 6 individuals undergoing total knee replacement for OA, were transduced with wild-type CBX4, mutant CBX4, and control lentiviral constructs. Senescence-related phenotypic outcomes included the following: multiple flow cytometry-measured markers (p16<jats:sup>INK4A<\/jats:sup>, senescence-associated \u03b2-galactosidase [SA-\u03b2-gal] activity and dipeptidyl peptidase-4 [DPP4], and proliferation marker EdU), multiplex ELISA-measured markers in chondrocyte culture media (senescence-associated secretory phenotypes [SASPs], including IL-1\u03b2, IL-6, IL-8, TNF-\u03b1, MMP-1, MMP-3, and MMP-9), and PCR array-evaluated senescence-related genes.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Results<\/jats:title>\n                <jats:p>Compared with control, CBX4 overexpression in OA chondrocytes decreased DPP4 expression and SASP secretion and increased chondrocyte proliferation confirming CBX4 senomorphic effects on primary human chondrocytes. Point mutations of the chromodomain domain (CDM, involved in chromatin modification) alone were sufficient to partially block the senomorphic activity of CBX4 (p16<jats:sup>INK4A<\/jats:sup> and DPP4 increased, and EdU decreased) but had minimal effect on SASP secretion. Although having no effect on p16<jats:sup>INK4A<\/jats:sup>, DPP4, and EdU, deletion of two small-ubiquitin-like-modifier-interaction motifs (CBX4 \u0394SIMs) led to increased SASP secretion (IL-1\u03b2, TNF-\u03b1, IL-8). The combination CBX4 CDM\u0394SIMs altered all these measures adversely and to a greater degree than the single domain mutants. Deletion of the C-terminal (CBX4 \u0394C-box) involved with transcriptional silencing of polycomb group proteins increased IL-1\u03b2 slightly but significantly but altered none of the other senescence outcome measures.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Conclusions<\/jats:title>\n                <jats:p>CBX4 has a senomorphic effect on human osteoarthritic chondrocytes. CDM is critical for CBX4-mediated regulation of senescence. The SIMs are supportive but not indispensable for CBX4 senomorphic function while the C-box is dispensable.<\/jats:p>\n              <\/jats:sec>","DOI":"10.1186\/s13075-023-03183-8","type":"journal-article","created":{"date-parts":[[2023,10,12]],"date-time":"2023-10-12T10:02:22Z","timestamp":1697104942000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Synergistic roles of CBX4 chromo and SIM domains in regulating senescence of primary human osteoarthritic chondrocytes"],"prefix":"10.1186","volume":"25","author":[{"given":"Yu-Hsiu","family":"Chen","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xin","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Attarian","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Virginia Byers","family":"Kraus","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2023,10,12]]},"reference":[{"key":"3183_CR1","doi-asserted-by":"crossref","unstructured":"Cui A, Li H, Wang D, Zhong J, Chen Y, Lu H. 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