Native and tagged CENP-A histones are functionally inequivalent.

IF 4.2 2区 生物学 Q1 GENETICS & HEREDITY
Minh Bui, Songjoon Baek, Reda S Bentahar, Daniël P Melters, Yamini Dalal
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引用次数: 0

Abstract

Background: Over the past several decades, the use of biochemical and fluorescent tags has elucidated mechanistic and cytological processes that would otherwise be impossible. The challenging nature of certain nuclear proteins includes low abundancy, poor antibody recognition, and transient dynamics. One approach to get around those issues is the addition of a peptide or larger protein tag to the target protein to improve enrichment, purification, and visualization. However, many of these studies were done under the assumption that tagged proteins can fully recapitulate native protein function.

Results: We report that when C-terminally TAP-tagged CENP-A histone variant is introduced, it undergoes altered kinetochore protein binding, differs in post-translational modifications (PTMs), utilizes histone chaperones that differ from that of native CENP-A, and can partially displace native CENP-A in human cells. Additionally, these tagged CENP-A-containing nucleosomes have reduced centromeric incorporation at early G1 phase and poorly associates with linker histone H1.5 compared to native CENP-A nucleosomes.

Conclusions: These data suggest expressing tagged versions of histone variant CENP-A may result in unexpected utilization of non-native pathways, thereby altering the biological function of the histone variant.

原生组蛋白和标记的 CENP-A 组蛋白在功能上是不等同的。
背景:在过去的几十年中,生化和荧光标签的使用阐明了原本不可能阐明的机理和细胞学过程。某些核蛋白的挑战性在于丰度低、抗体识别能力差以及瞬时动态性。解决这些问题的一种方法是在目标蛋白质上添加多肽或更大的蛋白质标签,以提高富集、纯化和可视化效果。然而,许多此类研究都是在假设标签蛋白能完全再现原生蛋白功能的前提下进行的:我们报告说,当引入 C 端 TAP 标记的 CENP-A 组蛋白变体时,它的动点核蛋白结合发生了改变,翻译后修饰(PTMs)不同,利用的组蛋白伴侣与原生 CENP-A 不同,并能在人体细胞中部分取代原生 CENP-A。此外,与原生 CENP-A 核小体相比,这些含有标记 CENP-A 的核小体在 G1 期早期的中心粒结合率较低,与连接组蛋白 H1.5 的结合率也较低:这些数据表明,表达标记版组蛋白变体 CENP-A 可能会导致非原生途径的意外利用,从而改变组蛋白变体的生物学功能。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Epigenetics & Chromatin
Epigenetics & Chromatin GENETICS & HEREDITY-
CiteScore
7.00
自引率
0.00%
发文量
35
审稿时长
1 months
期刊介绍: Epigenetics & Chromatin is a peer-reviewed, open access, online journal that publishes research, and reviews, providing novel insights into epigenetic inheritance and chromatin-based interactions. The journal aims to understand how gene and chromosomal elements are regulated and their activities maintained during processes such as cell division, differentiation and environmental alteration.
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