The sea cucumber Apostichopus japonicus requires its own defense mechanism to resist and adapt to high temperature stress. Epigenetic modification plays an important role in this regulation process. To investigate the changes of DNA methylation level and methylation pattern in A. japonicus (2-year-old) under different temperature stress, whole genomic bisulfite sequencing (WGBS) and enzyme-linked immunosorbent assay (ELISA) were used to detect whole genome methylation levels of the muscle, respiratory tree, digestive tract, and body wall. Three temperature gradients of 20℃, 26℃, and 32℃ were set up in this experiment. The results of WGBS showed that the total genome wide methylation levels of 20℃, 26℃, and 32℃ groups were (1.70±0.01)%, (1.79±0.11)%, and (1.59±0.04)%, respectively. The methylation level of the digestive tract genome of A. japonicus in the 26℃ group increased, while the methylation level of the 32℃ group decreased under high temperature stress. Among the total methylation sites, CG type was the main site of methylation modification (more than 96%), and CHH and CHG sites showed relatively low levels of modification. Among the methylation sites at 30% methylation level, CHG and CHH methylation sites showed the highest methylation, and were significantly higher than that of the CG type. The results of ELISA showed that the methylation levels of respiratory tree and digestive tract tissue ranged from 2.68% to 3.29% at three different temperatures, which were higher than those in muscle and body wall tissue. After temperature change, the total methylation level of respiratory tree and digestive tract tissue of sea cucumber changed significantly, while the total methylation level of muscle and body wall remained unchanged, indicating that DNA methylation may be involved in the regulation mechanism of high temperature stress in A. japonicus. The study on DNA methylation level of A. japonicus in response to temperature change can be used to analyze the effect of temperature rise on different tissues of sea cucumber from the perspective of epigenetics. This research can provide reliable basis for enriching the research content on methylation in A. japonicus and the occurrence of methylation in invertebrates.