Sousa-Canavez Juliana M, Beton Daniela, Gonzalez-Kristeller Daniela C, da Silva Aline M
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes 748, 05508-000, São Paulo, Brazil.
Biochimie. 2007 May;89(5):692-701. doi: 10.1016/j.biochi.2007.01.002. Epub 2007 Jan 21.
The protein phosphatase type-1 catalytic subunit (PP1c) does not exist freely in the cell and its activity must be very strictly controlled. Several protein inhibitors of PP1c have been described including the classical mammalian inhibitor-1 (I-1) and inhibitor-2 (I-2). Association of these inhibitors with PP1c appears to involve multiple contacts and in the case of I-2 no less than five I-2 interaction subdomains have been proposed. In this report, we provide both in vitro and in vivo evidence that the Dictyostelium discoideum genome encodes a protein (DdI-2) that is an ortholog of mammalian I-2, being the first PP1c interacting protein characterized in this social amoeba. Despite the low overall sequence similarity of DdI-2 with other I-2 sequences and its long N-terminal extension, the five PP1c interaction motifs proposed for mammalian I-2 are reasonably conserved in the Dictyostelium ortholog. We demonstrate that DdI-2 interacts with and inhibits D. discoideum PP1c (DdPP1c), which we have previously characterized. Moreover, using yeast two-hybrid assays we show that a stable interaction of DdI-2 with DdPP1c requires multiple contacts.
1型蛋白磷酸酶催化亚基（PP1c）在细胞内并非自由存在，其活性必须受到非常严格的控制。已经描述了几种PP1c的蛋白抑制剂，包括经典的哺乳动物抑制剂-1（I-1）和抑制剂-2（I-2）。这些抑制剂与PP1c的结合似乎涉及多个接触点，就I-2而言，已提出不少于五个I-2相互作用亚结构域。在本报告中，我们提供了体外和体内证据，表明盘基网柄菌基因组编码一种蛋白（DdI-2），它是哺乳动物I-2的直系同源物，是这种社会性变形虫中第一个被鉴定的与PP1c相互作用的蛋白。尽管DdI-2与其他I-2序列的总体序列相似性较低，且其N端延伸较长，但为哺乳动物I-2提出的五个PP-1c相互作用基序在盘基网柄菌直系同源物中相当保守。我们证明DdI-2与我们之前鉴定的盘基网柄菌PP1c（DdPP1c）相互作用并抑制其活性。此外，使用酵母双杂交试验我们表明DdI-2与DdPP1c的稳定相互作用需要多个接触点。
