Watanabe Mariko, Ogawa Yuji, Ito Kinji, Higashihara Masaaki, Kadin Marshall E, Abraham Lawrence J, Watanabe Toshiki, Horie Ryouichi
Fourth Department of Internal Medicine, Kitasato University School of Medicine, Kanagawa, Japan.
Am J Pathol. 2003 Aug;163(2):633-41. doi: 10.1016/S0002-9440(10)63690-5.
Overexpression of CD30 is the hallmark of Hodgkin and Reed-Sternberg (H-RS) cells and drives constitutive nuclear factor-kappaB activation that is the molecular basis for the pathophysiology of Hodgkin's lymphoma. Transcription of the CD30 gene is controlled by the core promoter that is driven by Sp-1 and the microsatellite sequences (MSs) that represses core promoter activity. To understand the mechanism(s) of CD30 overexpression in H-RS cells, we structurally and functionally characterized the CD30 MSs. Although the CD30 MS of H-RS cell lines was polymorphic, it was not truncated compared with that of control cells. A strong core promoter activity and constitutive Sp-1 binding were revealed in all cell lines examined irrespective of the levels of CD30 expression. In transient reporter gene assays, all MS clones derived from H-RS cell lines repressed the core promoter activity in unrelated cell lines, but not in the H-RS cell lines. An AP-1-binding site was found in the MS at nucleotide position of -377 to -371, the presence of which was found to relieve repression of the core promoter in H-RS cell lines but not in other tumor cell lines. H-RS cell lines showed constitutive and strong AP-1-binding activity, but other cell lines did not. The AP-1 complex contained JunB, whose overexpression activated reporter constructs driven by the CD30 promoter including the MSs, and was dependent on the AP-1 site. JunB expression was detected in H-RS cells in vitro and in vivo, but not in reactive cells or tumor cells of non-Hodgkin's lymphoma of diffuse large B-cell type. Transduction of JunB small interfering RNAs suppressed CD30 promoter activity in L428 cells but not in control cells. Taken together, overexpression and binding of JunB to the AP-1 site appear to relieve the repression of the core promoter by the CD30 MS in H-RS cells, which provide one basis for the constitutive overexpression of CD30 in Hodgkin's lymphoma.
CD30的过表达是霍奇金和里德-斯腾伯格（H-RS）细胞的标志，并驱动组成型核因子-κB激活，这是霍奇金淋巴瘤病理生理学的分子基础。CD30基因的转录受核心启动子控制，该核心启动子由Sp-1驱动，而微卫星序列（MSs）则抑制核心启动子活性。为了了解H-RS细胞中CD30过表达的机制，我们对CD30 MSs进行了结构和功能表征。尽管H-RS细胞系的CD30 MS是多态性的，但与对照细胞相比并未截短。在所检测的所有细胞系中均显示出强大的核心启动子活性和组成型Sp-1结合，而与CD30表达水平无关。在瞬时报告基因测定中，所有源自H-RS细胞系的MS克隆均抑制无关细胞系中的核心启动子活性，但在H-RS细胞系中则不然。在MS的核苷酸位置-377至-371处发现了一个AP-1结合位点，发现该位点的存在可减轻H-RS细胞系中核心启动子的抑制作用，但在其他肿瘤细胞系中则不然。H-RS细胞系显示出组成型且强大的AP-1结合活性，而其他细胞系则没有。AP-1复合物包含JunB，其过表达激活了由包括MSs的CD30启动子驱动的报告基因构建体，并且依赖于AP-1位点。在体外和体内的H-RS细胞中均检测到JunB表达，但在弥漫性大B细胞型非霍奇金淋巴瘤的反应性细胞或肿瘤细胞中未检测到。JunB小干扰RNA的转导抑制了L428细胞中的CD30启动子活性，但在对照细胞中则不然。综上所述，JunB的过表达及其与AP-1位点的结合似乎减轻了H-RS细胞中CD30 MS对核心启动子的抑制作用，这为霍奇金淋巴瘤中CD30的组成型过表达提供了一个基础。
