S-palmitoylation of protein is a posttranslational, reversible lipid modification; it was catalyzed by a family of 23 mammalian palmitoyl acyltransferases in humans. S-palmitoylation can impact protein function by regulating protein sorting, secretion, trafficking, stability, and protein interaction. Thus, S-palmitoylation plays a crucial role in many human diseases including mental illness and cancers. In this chapter, we systematically reviewed the influence of S-palmitoylation on protein performance, the characteristics of S-palmitoylation regulating protein function, and the role of S-palmitoylation in pulmonary inflammation and pulmonary hypertension and summed up the treatment strategies of S-palmitoylation-related diseases and the research status of targeted S-palmitoylation agonists/inhibitors. In conclusion, we highlighted the potential role of S-palmitoylation and depalmitoylation in the treatment of human diseases.