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bioRxiv · 10.64898/2026.10.23.761071

Tropomyosin 3.1 Regulates Chondrocyte Cortical F-actin Stability and Homeostasis

Herrick, M.·Arranguez, M.·Stack, J.·Parreno, J.
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Abstract

Objective: Osteoarthritis is a degenerative joint disease of which the mechanisms and proteins contributing to articular cartilage breakdown have not yet been fully elucidated. In osteoarthritis, chondrocyte F-actin loss coincides with altered chondrocyte homeostasis favoring catabolism. While the loss of F-actin has been implicated as a driver of osteoarthritis progression, the mechanisms by which F-actin is lost and regulates a catabolic shift in gene expression require elucidation. We hypothesized that destabilizing F-actin by targeting inhibition of specific F-actin stabilizer Tropomyosins (Tpms) dysregulates chondrocyte homeostasis through regulation of actin-associated transcription factors, Yes-associated protein (YAP) and Myocardin-related transcription factor A (MRTF). Methods: Using RT-PCR and Sanger sequencing, we determined the expression of Tpm isoforms present in cartilage. We then focused on the regulation of chondrocyte cortical F-actin by the specific Tpm isoform, Tpm3.1. Using confocal imaging for F-actin, we examined the effect of Tpm3.1 knockout in mouse femoral head cartilage and pharmacological inhibition (TR100) of primary bovine chondrocytes. Next, we examined the nuclear translocation of actin-associated regulators, YAP and MRTF, by TPM3.1 inhibition in primary chondrocytes. Finally, we determined the effect of YAP and MRTF inhibition on the regulation of chondrogenic and degradative enzyme gene expression in primary chondrocytes. Results: Our results revealed TPM3.1 is present and localized cortically in mouse and bovine chondrocytes. There was a reduction in cortical F-actin staining in Tpm3.1 knockout mouse femoral head cartilage as well as in primary bovine chondrocytes treated with TR100. In primary chondrocytes, TR100 caused a catabolic shift in mRNA levels. This coincided with a decrease in the nuclear-to-cytoplasmic ratio of both YAP and MRTF in TR100-treated cells. Furthermore, pharmacological inhibition of either YAP or MRTF partially recapitulated the catabolic shift seen in TR100-treated chondrocytes by a reduction of chondrogenic mRNA levels. Conclusion: Collectively, these results provide insight into the regulation of chondrocyte homeostasis by cortical F-actin stability and the downstream mediators of transcription. A greater understanding of the actin-associated molecules that regulate chondrocyte homeostasis in osteoarthritis may lead to novel insights to target disease progression.

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Herrick, M., Arranguez, M., Stack, J., Parreno, J.. 2026-10-26. Tropomyosin 3.1 Regulates Chondrocyte Cortical F-actin Stability and Homeostasis. https://doi.org/10.64898/2026.10.23.761071

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