Lehotzky Atilla, Tirián László, Tökési Natália, Lénárt Péter, Szabó Bálint, Kovács János, Ovádi Judit
Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, 1113 Budapest, Hungary.
J Cell Sci. 2004 Dec 1;117(Pt 25):6249-59. doi: 10.1242/jcs.01550.
Recently we identified TPPP/p25 (tubulin polymerization promoting protein/p25) as a brain-specific unstructured protein that induced aberrant microtubule assemblies and ultrastructure in vitro and as a new marker for Parkinson's disease and other synucleopathies. In this paper the structural and functional consequences of TPPP/p25 are characterized to elucidate the relationship between the in vitro and the pathological phenomena. We show that at low expression levels EGFP-TPPP/p25 specifically colocalizes with the microtubule network of HeLa and NRK cells. We found that the colocalization was dynamic (tg=5 seconds by fluorescence recovery after photobleaching) and changed during the phases of mitosis. Time-lapse and immunofluorescence experiments revealed that high levels of EGFP-TPPP/p25 inhibited cell division and promoted cell death. At high expression levels or in the presence of proteosome inhibitor, green fusion protein accumulated around centrosomes forming an aggresome-like structure protruding into the nucleus or a filamentous cage of microtubules surrounding the nucleus. These structures showed high resistance to vinblastin. We propose that a potential function of TPPP/p25 is the stabilization of physiological microtubular ultrastructures, however, its upregulation may directly or indirectly initiate the formation of aberrant protein aggregates such as pathological inclusions.
最近，我们鉴定出TPPP/p25（微管蛋白聚合促进蛋白/p25）是一种脑特异性无结构蛋白，它在体外可诱导异常微管组装和超微结构，并且是帕金森病和其他突触核蛋白病的一种新标志物。在本文中，对TPPP/p25的结构和功能后果进行了表征，以阐明体外现象与病理现象之间的关系。我们发现，在低表达水平时，EGFP-TPPP/p25与HeLa和NRK细胞的微管网络特异性共定位。我们发现这种共定位是动态的（光漂白后荧光恢复的半衰期为5秒），并且在有丝分裂阶段会发生变化。延时和免疫荧光实验表明，高水平的EGFP-TPPP/p25会抑制细胞分裂并促进细胞死亡。在高表达水平或存在蛋白酶体抑制剂的情况下，绿色融合蛋白在中心体周围积累，形成一种类似聚集体的结构，突出到细胞核中，或者形成围绕细胞核的微管丝状笼。这些结构对长春花碱具有高度抗性。我们提出，TPPP/p25的一个潜在功能是稳定生理性微管超微结构，然而，其上调可能直接或间接引发异常蛋白聚集体的形成，如病理性包涵体。
