Hypoxia-inducible factor 1α (HIF1α) is a critical transcriptional factor for inducing tumor metastasis, and stabilized under hypoxia but degraded by von Hippel–Lindau protein (pVHL) under normoxia. For the maximal degradation of HIF1α, it must be exported to the cytoplasm via an unidentified transporter. Here, we demonstrate that vitamin D3 up-regulated protein 1 (VDUP1) associates with the β-domain of pVHL and enhances the interaction between pVHL and HIF1α to promote the nuclear export and degradation of HIF1α hypoxia-independently. Blocking of VDUP1 translocation either by leptomycin B or by nuclear export signal mutation inhibited the nuclear export of pVHL/HIF1α and relieved the destabilization of HIF1α. VDUP1 suppressed cell invasiveness and tumor metastasis, which were also recovered by blocking of nuclear export. Taken together, these findings indicate that VDUP1 is a novel tumor suppressor which mediates the nuclear export of pVHL/HIF1α complex to destabilize HIF1α.