TPM3.1 association with actin stress fibers is required for lens epithelial to mesenchymal transition
| Autor: | Michael B. Amadeo, Velia M. Fowler, Elizabeth H. Kwon, Justin Parreno |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
Gene isoform
0303 health sciences Stress fiber Chemistry Tropomyosin Cell biology law.invention 03 medical and health sciences 0302 clinical medicine Downregulation and upregulation Cell culture Confocal microscopy law Epithelial–mesenchymal transition 030217 neurology & neurosurgery Actin 030304 developmental biology |
| DOI: | 10.1101/774794 |
| Popis: | PurposeEpithelial to mesenchymal transition (EMT) is a cause of anterior and posterior subcapsular cataracts. Central to EMT is the formation of actin stress fibers. Targeting specific, stress fiber associated tropomyosin in epithelial cells may be a means to prevent stress fiber formation and repress lens EMT.MethodsWe identified Tpm isoforms in mouse immortalized lens epithelial cells and isolated whole lenses by semi-quantitative PCR followed Sanger sequencing. We focused on the role of one particular tropomyosin isoform, Tpm3.1, in EMT. To stimulate EMT, we cultured cells or native lenses in TGFβ2. To test the function of Tpm3.1, we exposed cells or whole lenses to a Tpm3.1-specific chemical inhibitor, TR100, as well as investigated lenses from Tpm3.1 knockout mice. We examined stress fiber formation by confocal microscopy and assessed EMT progression by αsma mRNA (qPCR) and protein (WES immunoassay) analysis.ResultsLens epithelial cells express eight tropomyosin isoforms. Cell culture studies showed that TGFβ2 treatment results in an upregulation of Tpm3.1, which associates with actin in stress fibers. TR100 prevents stress fiber formation and reduces αsma in TGFβ2 treated cells. We confirmed the role of Tpm3.1 in lens epithelial cells in the native lens ex vivo. Culture of whole lenses in the presence of TGFβ2 results in stress fiber formation at the basal regions of epithelial cells. Knockout of Tpm3.1 or treatment of lenses with TR100 prevents basal stress fiber formation and reduces epithelial αsma levels.ConclusionTargeting specific stress fiber associated tropomyosin isoform, Tpm3.1, is a means to represses lens EMT. |
| Databáze: | OpenAIRE |
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