Ding Yuanying, Zhang Yong Z, Furuyama Kazumichi, Ogawa Kazuhiro, Igarashi Kazuhiko, Shibahara Shigeki
Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Sendai, Japan.
FEBS J. 2006 Dec;273(23):5333-46. doi: 10.1111/j.1742-4658.2006.05526.x. Epub 2006 Oct 25.
Intracellular heme concentrations are maintained in part by heme degradation, which is catalyzed by heme oxygenase. Heme oxygenase consists of two structurally related isozymes, HO-1 and HO-2. Recent studies have identified HO-2 as a potential oxygen sensor. To gain further insights into the regulatory role of HO-2 in heme homeostasis, we analyzed the expression profiles of HO-2 and the biochemical consequences of HO-2 knockdown with specific short interfering RNA (siRNA) in human cells. Both HO-2 mRNA and protein are expressed in the eight human cancer cell lines examined, and HO-1 expression is detectable in five of the cell lines, including HeLa cervical cancer and HepG2 hepatoma. Down-regulation of HO-2 expression with siRNA against HO-2 (siHO-2) caused induction of HO-1 expression at both mRNA and protein levels in HeLa and HepG2 cells. In contrast, knockdown of HO-1 expression did not noticeably influence HO-2 expression. HO-2 knockdown prolonged the half-life of HO-1 mRNA twofold in HeLa cells. Transient transfection assays in HeLa cells revealed that the 4.5-kb human HO-1 gene promoter was activated with selective knockdown of HO-2 in a sequence-dependent manner. Moreover, HO-2 knockdown caused heme accumulation in HeLa and HepG2 cells only when exposed to exogenous hemin. HO-2 knockdown may mimic a certain physiological change that is important in the maintenance of cellular heme homeostasis. These results suggest that HO-2 may down-regulate the expression of HO-1, thereby directing the co-ordinated expression of HO-1 and HO-2.
细胞内血红素浓度部分通过血红素降解来维持，血红素降解由血红素加氧酶催化。血红素加氧酶由两种结构相关的同工酶HO - 1和HO - 2组成。最近的研究已将HO - 2鉴定为一种潜在的氧传感器。为了进一步深入了解HO - 2在血红素稳态中的调节作用，我们分析了HO - 2的表达谱以及在人类细胞中用特异性短发夹RNA（siRNA）敲低HO - 2的生化后果。在所检测的8种人类癌细胞系中均表达HO - 2 mRNA和蛋白，并且在其中5种细胞系中可检测到HO - 1表达，包括HeLa宫颈癌和HepG2肝癌细胞系。用针对HO - 2的siRNA（siHO - 2）下调HO - 2表达导致HeLa和HepG2细胞中HO - 1在mRNA和蛋白水平上的表达诱导。相反，敲低HO - 1表达并未显著影响HO - 2表达。在HeLa细胞中，HO - 2敲低使HO - 1 mRNA的半衰期延长了两倍。HeLa细胞中的瞬时转染实验表明，4.5 kb的人类HO - 1基因启动子在HO - 2被选择性敲低时以序列依赖的方式被激活。此外，仅在暴露于外源性血红素时，HO - 2敲低才导致HeLa和HepG2细胞中的血红素积累。HO - 2敲低可能模拟了在维持细胞血红素稳态中重要的某种生理变化。这些结果表明，HO - 2可能下调HO - 1的表达，从而指导HO - 1和HO - 2的协调表达。
