人原肌球蛋白异构体比较的荧光蛋白标记。

IF 1.7 4区 生物学 Q3 BIOLOGY
Biology Open Pub Date : 2025-08-15 Epub Date: 2025-07-29 DOI:10.1242/bio.061992
Will Scott, Vitaliia Polutranko, Jakub Milczarek, Ian Hands-Portman, Mohan K Balasubramanian
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引用次数: 0

摘要

原肌球蛋白是一种重要的肌动蛋白细胞骨架蛋白,支持肌肉收缩、细胞形状和细胞分裂等过程。原肌球蛋白功能缺陷可导致疾病,包括一些肌病和过敏。在细胞中,原肌凝蛋白分子形成盘绕的二聚体,然后与其他二聚体端到端聚合,形成肌动蛋白结合。原肌球蛋白在不干扰其聚合界面的情况下标记体内荧光显微镜具有挑战性。我们最近开发了一种荧光标记,它包含一个40个氨基酸的柔性连接体,能够检测S. pombe肌动蛋白电缆和肌动球蛋白环中的原肌球蛋白,以及以前未被发现的斑块状结构。我们还使用这种策略成功地标记了人类TPM2.2,这是一种重要的人类肌肉异构体。在这里,我们扩展了这个工具,使用mNeonGreen、mCherry、mStayGold(E138D)和mScarlet3-H标签来可视化其他8种人类原肌球蛋白同工型。所有显示典型的原肌球蛋白荧光,没有细胞毒性的迹象,并与超分辨率显微镜兼容。这些工具单独或组合应该有助于详细的机制研究原肌球蛋白同型。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Fluorescent protein tags for human tropomyosin isoform comparison.

Tropomyosin is an important actin cytoskeletal protein underpinning processes such as muscle contraction, cell shape and cell division. Defects in tropomyosin function can lead to diseases, including some myopathies and allergies. In cells, tropomyosin molecules form coiled-coil dimers, which then polymerise end-to-end with other dimers for actin association. Tropomyosin is challenging to tag for in vivo fluorescence microscopy without perturbing its polymerisation interfaces. We recently developed a fluorescent tag comprising a 40-amino acid flexible linker capable of detecting tropomyosin in S. pombe actin cables and the actomyosin ring, and in patch-like structures that were previously unappreciated. We also used this strategy successfully to tag human TPM2.2, a prominent human muscle isoform. Here, we expanded this tool to visualise eight other human tropomyosin isoforms, using mNeonGreen, mCherry, mStayGold(E138D) and mScarlet3-H tags. All showed typical tropomyosin fluorescence, no signs of cytotoxicity and are compatible with super-resolution microscopy. These tools singly or in combination should aid detailed mechanistic investigations of tropomyosin isoforms.

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来源期刊
Biology Open
Biology Open BIOLOGY-
CiteScore
3.90
自引率
0.00%
发文量
162
审稿时长
8 weeks
期刊介绍: Biology Open (BiO) is an online Open Access journal that publishes peer-reviewed original research across all aspects of the biological sciences. BiO aims to provide rapid publication for scientifically sound observations and valid conclusions, without a requirement for perceived impact.
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