Here, we demonstrate that in human gastric cancer cells, the E3 ligase TRIM8 promotes the K63-linked ubiquitination of the glycolytic enzyme PGK1 and improves its stability, which leads to acetyltransferase ACAT1 recruitment, increased interaction of PGK1 with ACAT1, and subsequent PGK1 acetylation-dependent glycolytic activity. The gene discussed is ACAT1; the disease is gastric cancer.