Molecular cloning, characterization and isolation of novel spliced variants of the human ortholog of a rat estrogen-regulated membrane-associated protein, UO-44
Molecular cloning, characterization and isolation of novel spliced variants of the human ortholog of a rat estrogen-regulated membrane-associated protein, UO-44
复制标题
DOI：
10.1038/sj.onc.1207754
复制
发表时间：
2004-07-22
期刊：
影响因子：
8
通讯作者：
Huynh, H
中科院分区：
文献类型：
--
作者：
Leong, CTC;Ng, CY;Huynh, H
我们之前报道了雌激素调节的大鼠子宫-卵巢特异性互补DNA （UO-44）的表征。为了了解这种蛋白在人类卵巢癌和子宫癌的发生和发展中的作用，我们现在报道了人类同源物（HuUO-44）的克隆和表征。HuUO-44定位于染色体10q26.13，包含9个外显子。多个组织Northern blot在胰腺中检测到两个约2和3kb的HuU044转录本。RT-PCR结果表明，HuU044基因在2号外显子和6号外显子之间发生了一系列复杂的选择性剪接事件，产生了四种新的剪接变体：HuUO-44A、HuUO-44B、HuUO-44C和HuUO-44D。在HuU044中发现的推测功能基序是两个CUB结构域和一个透明带结构域。转染研究证实HuU044具有膜相关性。通过免疫组化，HuU044定位于正常卵巢和卵巢肿瘤上皮细胞；在NIH-OVCAR3型卵巢癌细胞中，HuU044仅在分裂细胞的边缘检测到。最重要的是，在多克隆HuU044抗血清存在下培养的NIH-OVCAR3细胞中，观察到细胞附着和增殖的明显丧失。这些发现提示HuU044在细胞运动、细胞-细胞基质中的潜在作用。
We have previously reported the characterization of an estrogen-regulated rat uterine-ovarian-specific complementary DNA (UO-44). To understand the involvement of this protein in the initiation and progression of human ovarian and uterine cancers, we now report the cloning and characterization of the human ortholog (HuUO-44). HuUO-44 is mapped to chromosome 10q26.13 and contains nine exons. Multiple tissue Northern blot detected two HuU044 transcripts of approximately 2 and 3kb in the pancreas. RT-PCR demonstrated that HuU044 undergoes a complex series of alternative splicing events between exons 2 and 6 that yielded four novel splice variants, HuUO-44A, HuUO-44B, HuUO-44C and HuUO-44D. Putative functional motifs identified in HuU044 are two CUB domains and a zona-pellucida domain. Transfection studies demonstrated the membrane-associated nature of HuU044. By immunohistochemistry, HuU044 was located to the normal ovarian and ovarian tumor epithelial cells; in NIH-OVCAR3 ovarian cancer cells, HuU044 was detected only at the leading edge of the dividing cells. Most importantly, a marked loss in cell attachment and proliferation was observed in NIH-OVCAR3 cells cultured in the presence of a polyclonal HuU044 antiserum. These findings suggest the potential role of HuU044 in cell motility, cell-cell matrices.