Pallier Coralie, Scaffidi Paola, Chopineau-Proust Stéphanie, Agresti Alessandra, Nordmann Patrice, Bianchi Marco E, Marechal Vincent
Unité de Virologie, Service de Bactériologie-Virologie, Hôpital de Bicêtre, Assistance Publique/Hôpitaux de Paris, 94275 Le Kremlin Bicêtre, France.
Mol Biol Cell. 2003 Aug;14(8):3414-26. doi: 10.1091/mbc.e02-09-0581. Epub 2003 Apr 17.
High mobility group box (HMGB) 1 and 2 are two abundant nonhistone nuclear proteins that have been found in association with chromatin. Previous studies based on immunofluorescence analysis indicated that HMGB1 dissociates from chromosomes during mitosis. In the present work, HMGB1 and 2 subcellular localization was reinvestigated in living cells by using enhanced green fluorescent protein- and Discosome sp. red fluorescent protein-tagged proteins. Contrary to previous reports, HMGB1 and 2 were shown to be present under two forms in mitotic cells, i.e., free and associated with the condensed chromatin, which rapidly exchange. A detailed analysis of HMGB2 interaction with mitotic chromosomes indicated that two sites encompassing HMG-box A and B are responsible for binding. Importantly, this interaction was rapidly inactivated when cells were permeabilized or exposed to chemical fixatives that are widely used in immunodetection techniques. A comparable behavior was also observed for two proteins of the HMG-nucleosome binding (HMGN) group, namely, HMGN1 and HMGN2.
高迁移率族蛋白盒（HMGB）1和2是两种在染色质中大量存在的非组蛋白核蛋白。以往基于免疫荧光分析的研究表明，HMGB1在有丝分裂期间从染色体上解离。在本研究中，通过使用增强型绿色荧光蛋白和盘状红荧光蛋白标记的蛋白，在活细胞中重新研究了HMGB1和2的亚细胞定位。与之前的报道相反，HMGB1和2在有丝分裂细胞中以两种形式存在，即游离形式和与浓缩染色质结合的形式，且它们能快速交换。对HMGB2与有丝分裂染色体相互作用的详细分析表明，包含HMG盒A和B的两个位点负责结合。重要的是，当细胞通透或暴露于免疫检测技术中广泛使用的化学固定剂时，这种相互作用会迅速失活。对于HMG-核小体结合（HMGN）组的两种蛋白，即HMGN1和HMGN2，也观察到了类似的行为。
