Molecular cloning, characterization, and temporal expression of the clock genes period and timeless in the oriental river prawn Macrobrachium nipponense during female reproductive development

The circadian clock is crucial for sustaining rhythmic biochemical, physiological, and behavioral processes in living creatures. In this study, we isolated and characterized two circadian clock genes in Macrobrachium nipponense, period (Mnper) and timeless (Mntim). The complete Mnper cDNA measures 4283 bp in length with an open reading frame encoding 1292 amino acids, including functional domains such as PER-ARNT-SIM (PAS), cytoplasmic localization domain (CLD), TIM interaction site (TIS), and nuclear localization signal (NLS). The deduced Mntim protein comprises1540 amino acids with functional domains such as PER interaction site (PIS), NLS, and CLD. Tissue distribution analyses showed that the two genes were highly expressed in the eyestalk and brain in both males and females, as well as being expressed in the ovary. The expression profiles of Mnper and Mntim were determined in the eyestalk, brain, and ovary under simulated breeding season and non-breeding season conditions. The expression profiles of both Mnper and Mntim appeared to be unaffected in the eyestalk. However, the expression of both genes exhibited significant seasonal variations in the brain, and thus we assumed the brain to be their functional location. The expression profiles under different simulated seasons and the variations during different ovarian stages indicate that both genes might be involved with female reproduction. Especially the mRNA levels in the brain varied greatly during these stages indicating that the clock function in the brain is closely related to ovarian development and female reproduction. And the reproductive roles of clock genes need to be elucidated.