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Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro
We previously discovered a cellular isoform of titin (originally named T-protein) colocalized with myosin II in the terminal web domain of the chicken intestinal epithelial cell brush border cytoskeleton (Eilertsen, K.J., and T.C.S. Keller. 1992. J. Cell Biol. 119:549-557). Here, we demonstrate that...
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Lenguaje: | English |
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The Rockefeller University Press
1994
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2120159/ https://www.ncbi.nlm.nih.gov/pubmed/8063857 |
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collection | PubMed |
description | We previously discovered a cellular isoform of titin (originally named T-protein) colocalized with myosin II in the terminal web domain of the chicken intestinal epithelial cell brush border cytoskeleton (Eilertsen, K.J., and T.C.S. Keller. 1992. J. Cell Biol. 119:549-557). Here, we demonstrate that cellular titin also colocalizes with myosin II filaments in stress fibers and organizes a similar array of myosin II filaments in vitro. To investigate interactions between cellular titin and myosin in vitro, we purified both proteins from isolated intestinal epithelial cell brush borders by a combination of gel filtration and hydroxyapatite column chromatography. Electron microscopy of brush border myosin bipolar filaments assembled in the presence and absence of cellular titin revealed a cellular titin- dependent side-by-side and end-to-end alignment of the filaments into highly ordered arrays. Immunogold labeling confirmed cellular titin association with the filament arrays. Under similar assembly conditions, purified chicken pectoralis muscle titin formed much less regular aggregates of muscle myosin bipolar filaments. Sucrose density gradient analyses of both cellular and muscle titin-myosin supramolecular arrays demonstrated that the cellular titin and myosin isoforms coassembled with a myosin/titin ratio of approximately 25:1, whereas the muscle isoforms coassembled with a myosin:titin ratio of approximately 38:1. No coassembly aggregates were found when cellular myosin was assembled in the presence of muscle titin or when muscle myosin was assembled in the presence of cellular titin. Our results demonstrate that cellular titin can organize an isoform-specific association of myosin II bipolar filaments and support the possibility that cellular titin is a key organizing component of the brush border and other myosin II-containing cytoskeletal structures including stress fibers. |
format | Text |
id | pubmed-2120159 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1994 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21201592008-05-01 Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro J Cell Biol Articles We previously discovered a cellular isoform of titin (originally named T-protein) colocalized with myosin II in the terminal web domain of the chicken intestinal epithelial cell brush border cytoskeleton (Eilertsen, K.J., and T.C.S. Keller. 1992. J. Cell Biol. 119:549-557). Here, we demonstrate that cellular titin also colocalizes with myosin II filaments in stress fibers and organizes a similar array of myosin II filaments in vitro. To investigate interactions between cellular titin and myosin in vitro, we purified both proteins from isolated intestinal epithelial cell brush borders by a combination of gel filtration and hydroxyapatite column chromatography. Electron microscopy of brush border myosin bipolar filaments assembled in the presence and absence of cellular titin revealed a cellular titin- dependent side-by-side and end-to-end alignment of the filaments into highly ordered arrays. Immunogold labeling confirmed cellular titin association with the filament arrays. Under similar assembly conditions, purified chicken pectoralis muscle titin formed much less regular aggregates of muscle myosin bipolar filaments. Sucrose density gradient analyses of both cellular and muscle titin-myosin supramolecular arrays demonstrated that the cellular titin and myosin isoforms coassembled with a myosin/titin ratio of approximately 25:1, whereas the muscle isoforms coassembled with a myosin:titin ratio of approximately 38:1. No coassembly aggregates were found when cellular myosin was assembled in the presence of muscle titin or when muscle myosin was assembled in the presence of cellular titin. Our results demonstrate that cellular titin can organize an isoform-specific association of myosin II bipolar filaments and support the possibility that cellular titin is a key organizing component of the brush border and other myosin II-containing cytoskeletal structures including stress fibers. The Rockefeller University Press 1994-09-01 /pmc/articles/PMC2120159/ /pubmed/8063857 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/). |
spellingShingle | Articles Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title | Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title_full | Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title_fullStr | Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title_full_unstemmed | Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title_short | Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro |
title_sort | cellular titin localization in stress fibers and interaction with myosin ii filaments in vitro |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2120159/ https://www.ncbi.nlm.nih.gov/pubmed/8063857 |