NF-κB mediates aberrant activation of HIF-1 in malignant lymphoma.

ABSTRACT

OBJECTIVE: The goal of this study was to explore the molecular mechanisms of aberrant hypoxia inducible factor-1 (HIF-1) activation in lymphoma cells. MATERIALS AND METHODS: We analyzed the expression of the α subunit of HIF-1 in three lymphoma cell lines and in normal CD19-positive B cells by Western blotting. To investigate the role of nuclear factor (NF)-κB in abnormal HIF-1α expression in lymphoma cells, we performed a reporter assay using HIF-1α promoter constructs that contained or lacked an NF-κB binding site. We also used a chromatin immunoprecipitation assay to assess whether NF-κB binds the HIF-1α promoter. In addition, we took advantage of NF-κB inhibitors. To analyze the function of HIF-1 in lymphoma cells, we established stable HIF-1α knockdown cells using short-hairpin RNA. RESULTS: Malignant lymphoma cells, but not normal B cells, demonstrated constitutive expression of HIF-1α. Inhibitors of NF-κB, however, drastically suppressed this HIF-1α expression at both the messenger RNA and protein levels. Furthermore, we found that exposure of lymphoma cells to ionizing radiation clearly induced NF-κB activation and increased HIF-1α expression. Suppressing HIF-1α expression by short-hairpin RNA increased the sensitivity of lymphoma cells to ionizing radiation-induced cell death. In searching for downstream targets of the NF-κB/HIF-1 axis, we identified survivin, a member of the IAP family of anti-apoptotic proteins. CONCLUSIONS: We found that aberrant activation of HIF-1 in malignant lymphoma cells was mediated, at least in part, by NF-κB activity. Our observations suggest that HIF-1 inhibition may be an effective strategy to improve the outcomes of lymphoma patients treated with radiation.