CHARACTERIZATION OF ENDOGENOUS HYPDXIA-INDUCED TRANSCRIPTIONAL REPRESSOR HIF-3ALPHA

缺氧诱导因子（hypoxia -inducible factor， HIF）是一种重要的转录因子，在哺乳动物细胞缺氧时被激活，这是一种肿瘤微环境条件，在肿瘤的进展和治疗中起着关键作用。HIF由氧敏感的HIF-α和组成性表达的HIF-β亚基组成。最近发现的人类HIF-α的第三个成员HIF-3α产生多个剪接变体。本文从人肾中克隆了HIF-3α剪接变体HIF-3α2的cDNA，并对其特征进行了分析。HIF-3α2以及首次发现的HIF-3α变体HIF-3α1在COS-7细胞中异位表达时，定位于细胞核，在缺氧条件下稳定表达。然而，报告基因分析表明，HIF-3α2不能激活hif介导的转录，相反，HIF-3a1具有与HIF-3α2相同的n端结构域，能够激活HIF-3α2。c端缺失分析显示HIF-3α2的c端片段掩盖了其自身的转录活性。我们进一步发现HIF-3α2在肾癌VMRC中的过表达抑制了缺氧诱导基因的体外表达和体内异种移植物的生长。这些结果表明，HIF-3α2在低氧条件下处于稳定状态，并以一种新的反馈机制对低氧诱导基因的表达起负调控作用。

Hypoxia-inducible factor (HIF) is an important transcriptional factor that is activated when mammalian cells experience hypoxia, a tumor microenvironmental condition that plays a pivotal role in tumor progression and treatment. HIF consists of oxygen-sensitive HIF-α and constitutively expressed HIF-β subunits. The recently identified third member of the human HIF-α, HIF-3α, produces multiple splicing variants. Here we cloned cDNA for one of the splicing variants of human HIF-3α, HIF-3α2, from human kidney and examined its characteristics. HIF-3α2, as well as the first identified HIF-3α variant HIF-3α1, localized in the nucleus and was stabilized under hypoxia when ectopically expressed in COS-7 cells. However, reporter gene analysis showed that HIF-3α2 failed to activate HIF-mediated transcription and conversely suppressed it, although HIF-3a1 with an N-terminal domain identical to HIF-3α2 had the ability to activate it. C-terminal deletion analysis revealed that the C-terminal moiety of HIF-3α2 masked its own transcriptional activity. We further showed that overexpression of HIF-3α2 in renal carcinoma VMRC inhibited both hypoxia-inducible gene expression in vitro and growth as xenografts in vivo. These results indicated that HIF-3α2, which is stabilized under hypoxia, functions as a negative regulator of hypoxia-inducible gene expression in a novel feedback mechanism.