The cellular prion protein (PrP) is associated with metastasis, tumor progression and recurrence; however, the precise mechanisms underlying its action is not well understood. Our study found that PrP degradation decreased tumor progression in colorectal cancer (CRC). In a CRC cell line and human CRC tissue exposed to hypoxia, induced heat-shock 70-kDa protein-1-like (HSPA1L) expression stabilized hypoxia-inducible factor-1α (HIF-1α) protein and promoted PrP accumulation and tumorigenicity in vivo. PrP was degraded via the proteasome pathway mediated by the ubiquitin-protein E3 ligase glycoprotein 78 (GP78), which interacts directly with PrP. However, hypoxia-induced HSPA1L interacted with GP78 and inhibited its functions. HSPA1L knockdown facilitated the interaction of GP78 and PrP, thereby increasing PrP ubiquitination. Thus, GP78 was identified as the ubiquitinase for PrP, thereby revealing an essential mechanism that controls PrP levels in CRC. Our results suggest that the HSPA1L/HIF-1α/GP78 axis has a crucial role in PrP accumulation during tumor progression.