All living organisms are influenced by the circadian clock. This mechanism generates rhythms that affect organismal processes involved in cellular activity, behavior and immunity. In the present study we investigated the function of the circadian clock genes: cry1a and cry1b in zebrafish larvae, focusing on their roles in early development, behavior, microbiota composition, and antiviral immune response. Using Clustered Regularly Interspaced Short Palindromic Repeats - CRISPR-associated protein 9 (CRISPR-Cas9) gene editing, we generated zebrafish lines with targeted mutations in cry1a and cry1b genes and found that cry1 genes are not essential for larval survival or development. However, both mutations led to significant behavioral changes, with mutant larvae exhibiting increased locomotor activity compared to their wild type (WT) counterparts. Microbiome analysis revealed that cry1a mutation, in particular, resulted in a higher abundance of Proteobacteria in the larval microbiota. When challenged with Tilapia lake virus (TiLV), both cry1a and cry1b mutants displayed increased viral loads and reduced survival rates, indicating a compromised antiviral response. Specifically, cry1a mutants had lower initial expression of tlr22, while cry1b mutants showed reduced gene expression of antiviral Mxa at later stages post-infection, both contributing to decreased resistance against TiLV. Overall, the findings demonstrate that while cry1a and cry1b mutations do not significantly affect an early development, both genes play important roles in regulating behavior, shaping the microbiome, and modulating innate antiviral immunity in zebrafish larvae. The study highlights the multifaceted functions of circadian clock genes in vertebrate physiology and immunity.
Circadian Biology