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Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition
PURPOSE: Epithelial to mesenchymal transition (EMT) is a cause of anterior and posterior subcapsular cataracts. Central to EMT is the formation of actin stress fibers. Selective targeting of actin stress fiber-associated tropomyosin (Tpm) in epithelial cells may be a means to prevent stress fiber fo...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Association for Research in Vision and Ophthalmology
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415280/ https://www.ncbi.nlm.nih.gov/pubmed/32492110 http://dx.doi.org/10.1167/iovs.61.6.2 |
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author | Parreno, Justin Amadeo, Michael B. Kwon, Elizabeth H. Fowler, Velia M. |
author_facet | Parreno, Justin Amadeo, Michael B. Kwon, Elizabeth H. Fowler, Velia M. |
author_sort | Parreno, Justin |
collection | PubMed |
description | PURPOSE: Epithelial to mesenchymal transition (EMT) is a cause of anterior and posterior subcapsular cataracts. Central to EMT is the formation of actin stress fibers. Selective targeting of actin stress fiber-associated tropomyosin (Tpm) in epithelial cells may be a means to prevent stress fiber formation and repress lens EMT. METHODS: We identified Tpm isoforms in mouse immortalized lens epithelial cells and epithelial and fiber cells from whole lenses by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) followed Sanger sequencing. We focused on the role of one particular tropomyosin isoform, Tpm3.1, in EMT. To induce EMT, we treated cells or native lenses with 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 analysis of alpha-smooth muscle actin (αSMA) mRNA (real-time RT-PCR), and protein (Western immunoassay [WES]). RESULTS: Lens epithelial cells express eight Tpm isoforms. Cell culture studies showed that TGFβ2 treatment results in the upregulation of Tpm3.1, which associates with actin in stress fibers. TR100 prevents stress fiber formation and reduces αSMA in TGFβ2-treated cells. Using an ex vivo lens culture model, TGFβ2 treatment results in stress fiber formation at the basal regions of the epithelial cells. Genetic knockout of Tpm3.1 or treatment of lenses with TR100 prevents basal stress fiber formation and reduces epithelial αSMA levels. CONCLUSIONS: Targeting specific stress fiber associated tropomyosin isoform, Tpm3.1, is a means to repress lens EMT. |
format | Online Article Text |
id | pubmed-7415280 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | The Association for Research in Vision and Ophthalmology |
record_format | MEDLINE/PubMed |
spelling | pubmed-74152802020-08-24 Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition Parreno, Justin Amadeo, Michael B. Kwon, Elizabeth H. Fowler, Velia M. Invest Ophthalmol Vis Sci Lens PURPOSE: Epithelial to mesenchymal transition (EMT) is a cause of anterior and posterior subcapsular cataracts. Central to EMT is the formation of actin stress fibers. Selective targeting of actin stress fiber-associated tropomyosin (Tpm) in epithelial cells may be a means to prevent stress fiber formation and repress lens EMT. METHODS: We identified Tpm isoforms in mouse immortalized lens epithelial cells and epithelial and fiber cells from whole lenses by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) followed Sanger sequencing. We focused on the role of one particular tropomyosin isoform, Tpm3.1, in EMT. To induce EMT, we treated cells or native lenses with 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 analysis of alpha-smooth muscle actin (αSMA) mRNA (real-time RT-PCR), and protein (Western immunoassay [WES]). RESULTS: Lens epithelial cells express eight Tpm isoforms. Cell culture studies showed that TGFβ2 treatment results in the upregulation of Tpm3.1, which associates with actin in stress fibers. TR100 prevents stress fiber formation and reduces αSMA in TGFβ2-treated cells. Using an ex vivo lens culture model, TGFβ2 treatment results in stress fiber formation at the basal regions of the epithelial cells. Genetic knockout of Tpm3.1 or treatment of lenses with TR100 prevents basal stress fiber formation and reduces epithelial αSMA levels. CONCLUSIONS: Targeting specific stress fiber associated tropomyosin isoform, Tpm3.1, is a means to repress lens EMT. The Association for Research in Vision and Ophthalmology 2020-06-03 /pmc/articles/PMC7415280/ /pubmed/32492110 http://dx.doi.org/10.1167/iovs.61.6.2 Text en Copyright 2020 The Authors http://creativecommons.org/licenses/by-nc-nd/4.0/ This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License. |
spellingShingle | Lens Parreno, Justin Amadeo, Michael B. Kwon, Elizabeth H. Fowler, Velia M. Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title | Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title_full | Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title_fullStr | Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title_full_unstemmed | Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title_short | Tropomyosin 3.1 Association With Actin Stress Fibers is Required for Lens Epithelial to Mesenchymal Transition |
title_sort | tropomyosin 3.1 association with actin stress fibers is required for lens epithelial to mesenchymal transition |
topic | Lens |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415280/ https://www.ncbi.nlm.nih.gov/pubmed/32492110 http://dx.doi.org/10.1167/iovs.61.6.2 |
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