Genomic variation in the SKA2 gene has recently been identified as a promising suicide biomarker. In light of its role in glucocorticoid receptor transactivation, we investigated whether SKA2 DNA methylation influences cortisol stress reactivity and is involved in the development of post-traumatic stress disorder (PTSD). Increased SKA2 methylation was significantly associated with lower cortisol stress reactivity in 85 healthy individuals exposed to the Trier Social Stress Test (B = -173.40, t = -2.324, p-value = 0.023). Next, we observed that longitudinal decreases in SKA2 methylation after deployment were associated with the emergence of post-deployment PTSD symptoms in a Dutch military cohort (N = 93; B = -0.054, t = -3.706, p-value = 3.66 x 10(-4)). In contrast, exposure to traumatic stress during deployment by itself resulted in longitudinal increases in SKA2 methylation (B = 0.037, t = 4.173, p-value = 6.98 x 10(-5)). Using predeployment SKA2 methylation levels and childhood trauma exposure, we found that the previously published suicide prediction rule significantly predicted post-deployment PTSD symptoms (AUC = 0.66, 95% CI: 0.53-0.79) with an optimal sensitivity of 0.81 and specificity of 0.91. Permutation analysis using random methylation loci supported these findings. Together, these data establish the importance of SKA2 for cortisol stress responsivity and the development of PTSD and provide further evidence that SKA2 is a promising biomarker for stress-related disorders including PTSD.