Entamoeba histolytica expressing a dominant negative N-truncated light subunit of its Gal-Lectin are less virulent
Entamoeba histolytica expressing a dominant negative N-truncated light subunit of its Gal-Lectin are less virulent
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DOI：
10.1091/mbc.e02-06-0344
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发表时间：
2002-12-01
影响因子：
3.3
通讯作者：
Mirelman, D
中科院分区：
文献类型：
--
作者：
Katz, U;Ankri, S;Mirelman, D
溶组织内阿米巴的 260 kDa 异二聚体 Gal/GalNAc 特异性凝集素（Gal-凝集素）在还原条件下解离成重亚基（hgl，170 kDa）和轻亚基（lgl，35 kDa）。我们之前已经证明，反义 RNA 抑制 35 kDa 亚基的表达会导致毒力降低。为了进一步了解半乳糖凝集素轻亚基在发病机制中的作用，用编码完整、突变和截短形式的轻亚基lgl1基因的质粒转染变形虫。其中lgl的55个N-末端氨基酸被去除的转染子，过量产生N-截短的lgl蛋白（32kDa），其在半乳糖凝集素异二聚体复合物的形成中取代了大部分天然35-kDalgl，并产生显性负效应，用该构建体转染的阿米巴显示其粘附和杀死哺乳动物细胞的能力以及与哺乳动物细胞形成玫瑰花结的能力显着下降。吞噬红细胞。此外，这种带有抗半乳糖凝集素抗体的转染子的免疫荧光共聚焦显微镜显示，加帽能力受损。这些结果表明，轻亚基在半乳糖凝集素复合物的聚集中发挥作用，并且其 N 截短影响该功能，这是毒力所必需的。
The 260-kDa heterodimeric Gal/GalNAc-specific Lectin (Gal-lectin) of Entamoeba histolytica dissociates under reducing conditions into a heavy (hgl, 170 kDa.) and a light subunit (lgl, 35 kDa). We have previously shown that inhibition of expression of the 35-kDa subunit by antisense RNA causes a decrease in virulence. To further understand the role of the light subunit of the Gal-lectin in pathogenesis, amoebae were transfected with plasmids encoding intact, mutated, and truncated forms of the light subunit lgl1 gene. A transfectant in which the 55 N-terminal amino acids of the lgl were removed, overproduced an N-truncated lgl protein (32 kDa), which replaced most of the native 35-kDa lgl in the formation of the Gal-lectin heterodimeric complex and exerted a dominant negative effect Amoebae transfected with this construct showed a significant decrease in their ability to adhere to and kill mammalian cells as well as in their capacity to form rosettes with and to plagocytose erythrocytes. In addition, immunofluorescence confocal microscopy of this transfectant with anti-Gal-lectin anti-bodies showed an impaired ability to cap. These results indicate that the light subunit has a role in enabling the clustering of Gal-lectin complexes and that its N-truncation affects this function, which is required for virulence.