Carlo Joelle M, Osman Ahmed, Niles Edward G, Wu Wenjie, Fantappie Marcelo R, Oliveira Francisco M B, LoVerde Philip T
Department of Microbiology and Immunology, School of Medicine and Biomedical Sciences, State University of New York, NY, USA.
FEBS J. 2007 Aug;274(16):4075-93. doi: 10.1111/j.1742-4658.2007.05930.x. Epub 2007 Jul 16.
Smad proteins are the cellular mediators of the transforming growth factor-beta superfamily signals. Herein, we describe the isolation of a fourth Smad gene from the helminth Schistosoma mansoni, a receptor-regulated Smad (R-Smad) gene termed SmSmad1B. The SmSmad1B protein is composed of 380 amino acids, and contains conserved MH1 and MH2 domains separated by a short 42 amino acid linker region. The SmSmad1B gene (> 10.7 kb) is composed of five exons separated by four introns. On the basis of phylogenetic analysis, SmSmad1B demonstrates homology to Smad proteins involved in the bone morphogenetic protein pathway. SmSmad1B transcript is expressed in all stages of schistosome development, and exhibits the highest expression level in the cercariae stage. By immunolocalization experiments, the SmSmad1B protein was detected in the cells of the parenchyma of adult schistosomes as well as in female reproductive tissues. Yeast two-hybrid experiments revealed an interaction between SmSmad1B and the common Smad, SmSmad4. As determined by yeast three-hybrid assays and pull-down assays, the presence of the wild-type or mutated SmTbetaRI receptor resulted in a decreased interaction between SmSmad1B and SmSmad4. These results suggest the presence of a nonfunctional interaction between SmSmad1B and SmTbetaRI that does not give rise to the phosphorylation and the release of SmSmad1B to form a heterodimer with SmSmad4. SmSmad1B, as well as the schistosome bone morphogenetic protein-related Smad SmSmad1 and the transforming growth factor-beta-related SmSmad2, interacted with the schistosome coactivator proteins SmGCN5 and SmCBP1 in pull-down assays. In all, these data suggest the involvement of SmSmad1B in critical biological processes such as schistosome reproductive development.
Smad蛋白是转化生长因子-β超家族信号的细胞介质。在此，我们描述了从曼氏血吸虫这种蠕虫中分离出的第四个Smad基因，这是一个名为SmSmad1B的受体调节型Smad（R-Smad）基因。SmSmad1B蛋白由380个氨基酸组成，包含保守的MH1和MH2结构域，中间由一个短的42个氨基酸的连接区隔开。SmSmad1B基因（>10.7 kb）由五个外显子和四个内含子组成。基于系统发育分析，SmSmad1B与骨形态发生蛋白途径中涉及的Smad蛋白具有同源性。SmSmad1B转录本在血吸虫发育的所有阶段均有表达，在尾蚴阶段表达水平最高。通过免疫定位实验，在成年血吸虫实质细胞以及雌性生殖组织中检测到了SmSmad1B蛋白。酵母双杂交实验揭示了SmSmad1B与普通Smad蛋白SmSmad4之间存在相互作用。通过酵母三杂交分析和下拉分析确定，野生型或突变型SmTbetaRI受体的存在导致SmSmad1B与SmSmad4之间的相互作用减弱。这些结果表明SmSmad1B与SmTbetaRI之间存在无功能的相互作用，这种相互作用不会导致SmSmad1B的磷酸化和释放，从而无法与SmSmad4形成异二聚体。在下拉分析中，SmSmad1B以及与血吸虫骨形态发生蛋白相关的Smad蛋白SmSmad1和与转化生长因子-β相关的SmSmad2，均与血吸虫共激活蛋白SmGCN5和SmCBP1相互作用。总之，这些数据表明SmSmad1B参与了诸如血吸虫生殖发育等关键生物学过程。
