Histone acetylation is a dynamic epigenetic modification and sensitive to the changes in extracellular environment. Butyrate, a histone deacetylase inhibitor, can inhibit the deacetylation process of histones. In this study, we found that the acetylation level of H3 was enhanced at 12 h after lipopolysaccharide (LPS) stimulation and increased at 6 h after combining treatment with LPS and butyrate in pearl oyster Pinctada fucata martensii. Transcriptome analysis indicated that butyrate counter-regulated 29.95%–36.35% of the genes repressed by LPS, and these genes were mainly enriched in the “cell proliferation” and “Notch signaling pathway”. Meanwhile, butyrate inhibited the up-regulation of 31.54%–54.96% of the genes induced by LPS, and these genes were mainly enriched in “Notch signaling pathway”, “cell proliferation”, “NF-kappa B signaling pathway”, “TNF signaling pathway”, “apoptosis”, “NOD-like receptor signaling pathway”, “RIG-I-like receptor signaling pathway” and “cytosolic DNA-sensing pathway”. Gene expression analysis showed that butyrate downregulated most of cell proliferation, immune-related genes effected by LPS. The activities of LAP, LYS, ACP, ALP, and GSH-Px were up-regulated at 6 h after combining treatment with LPS and butyrate, suggesting that butyrate could activate serum immune-related enzymes in pearl oyster. These results can improve our understanding of the function of histone deacetylase in the immune response of pearl oyster and provide references for an in-depth study of the functions of histone deacetylase in mollusks.