Arrest-defective-1 protein, an acetyltransferase, does not alter stability of hypoxia-inducible factor (HIF)-1alpha and is not induced by hypoxia or HIF.

The hypoxia-inducible factor (HIF) is a key player in a transcriptional pathway that controls the hypoxic response of mammalian cells. Post-translational modification of the alpha subunit of HIF determines its half-life and activity. Among the multiple reported modifications, acetylation, by an acetyltransferase termed arrest-defective-1 protein (ARD1), has been reported to decrease HIF-1alpha stability and therefore impact on hypoxic gene expression. In contrast, we report that both overexpression and silencing of ARD1 had no impact on the stability of HIF-1alpha or -2alpha and that cells silenced for ARD1 maintained hypoxic nuclear localization of HIF-1alpha. In addition, we show that the ARD1 mRNA and protein levels are not regulated by hypoxia in several human tumor cell lines, including cervical adenocarcinoma HeLa cells, fibrosarcoma HT1080 cells, adenovirus-transformed human kidney HEK293 cells, and human breast cancer MCF-7 cells. Using two model systems ((a) wild-type and HIF-1alpha-null mouse embryo fibroblasts and (b) HeLa cells silenced for HIF-1alpha or -2alpha by RNA interference), we demonstrate that the level of expression of the ARD1 protein is independent of HIF-1alpha and -2alpha. We also demonstrate that ARD1 is a stable, predominantly cytoplasmic protein expressed in a broad range of tissues, tumor cell lines, and endothelial cells. Taken together, our findings demonstrate that ARD1 has limited, if any, impact on the HIF signaling pathway.