Allene oxide synthase (AOS) and hydroperoxide lyase (HPL), members of the CYP74 gene family, are branches of the oxylipin pathway and play vital roles in plant responses to a number of stresses. In this study, four HPL genes and one AOS gene were identified in the watermelon genome, which were clustered into three subfamilies (CYP74A, CYP74B and CYP74C). Sequence analysis revealed that most HPL and AOS proteins from various plants contain representative domains, including Helix-I region, Helix-K region (ExxR) and Heme-binding domain. A number of development-, stress-, and hormone-related cis-elements were found in the promoter regions of the ClAOS and ClHPL genes, and the detected ClAOS and ClHPL genes were differentially expressed in different tissues and fruit development stages, as well as in response to various hormones. In addition, red light could enhance the expression of ClAOS in root-knot nematode-infected leaves and roots of watermelon, implying that ClAOS might play a primary role in red light-induced resistance against root-knot nematodes. These findings lay a foundation for understanding the specific function of CYP74 genes in watermelon.Agronomy 2019, 9, 872 2 of 17 isomerization or dehydration of fatty acid hydroperoxides and require neither molecular oxygen nor NAD(P)H-dependent cytochrome P450-reductase [1,2,6].CYP74 enzymes are broadly classified into four subfamilies (CYP74A, CYP74B, CYP74C, and CYP74D) based on their evolutionary heritage and relationship with the diversity of oxylipin structure [6]. AOS and DES members constitute the CYP74A and CYP74D subfamily, respectively, while CYP74B and CYP74C subfamilies contain enzymes with HPL activity [5,7]. In addition, AOS enzymes can be classified into three different types: The first two types can use either 13-hydroperoxide derivatives or 9-hydroperoxide derivatives as the substrate (13-AOS and 9-AOS, respectively), while the third type can use both of them (9/13-AOS) as the substrate [4,8]. Similarly, three types of HPL enzymes named as 13-HPL, 9-HPL, and 9/13-HPL were also reported in a number of plant species [9]. To date, AOS and HPL genes have been cloned and functionally characterized from a number of plant species, such as tomato (Solanum lycopersicum L. (Solanaceae)) [10,11], tobacco (Nicotiana attenuate Torr. ex S.Watson) [12], rice (Oryza sativa L.) [13,14], and barrel medic (Medicago truncatula Gaertn.) [15], whereas DES genes have been isolated in very few plants [16][17][18][19][20]. These findings reveal that plant CYP74 family genes are indispensable for oxylipin biosynthesis, and play important roles in plant growth and development, as well as in plant defense to various abiotic and biotic stresses.Watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) is an agricultural crop with high nutritional and economic values, but it is rather susceptible to various biotic and abiotic stresses during growth and development [21]. The root-knot nematodes (RKNs, Meloidogyne incognita) mainly infect the roots of host plants and increase sus...