Regulatory Genes in Eyespot Formation and Function of Mytilus coruscus

Light sensitivity is important for marine benthic invertebrates, and it plays a vital role in the marine bivalves settling. Animal visual systems are enormously diverse; their development appears to be controlled by a set of conserved retinal determination genes (RDGs). Eyespots, as the simplest animal eyes, their appearance indicates the important effect on mussel larvae attachment. Nevertheless, the molecular mechanism of the eyespot's development in Mytilus coruscus larvae is not clear. In this study, we identified 11 genes which play a regulatory role in the visual system (i.e. Pax1/9, Pax2/5/8, Pax6, Pax3/7, Six1/2, Six3/6, Six4/5, Dach, Eya, Brn and Tbx2) from transcriptome data and the whole genome sequence of M. coruscus. The results of chromosome localization showed that 11 genes were distributed on different chromosomes. Subcellular mapping revealed that all the proteins except Brn were located in the nucleus. Phylogeny and gene structure analyses revealed that the Pax members were divided into four subfamilies, the Six members were divided into three subfamilies and structures within the same subfamily were relatively conserved. Quantitative real-time PCR (qPCR) showed that Dach, Pax6, Pax3/7, Six1/2 and Six4/5 were expressed at high levels during the pediveliger stage. Moreover, Six1/2 and Six4/5 were highly expressed in mantle tissues. Subsequent overall in situ hybridization experiments in the planktonic larval stage revealed that Pax6, Six1/2 and Six4/5 detected signals in the region of the eyespot. Based on these analyses, we suggested that the development of vision in M. coruscus not only depended on the expression pattern of Pax6, but perhaps also related to Six1/2 and Six4/5 in the planktonic larval stage, while Six1/2 and Six4/5 were the dominant genes for visual function in the adult mussel. This study made a comprehensive analysis of the visual function of M. coruscus at the genome level, which helps us to understand the intrinsic mechanism of the visual system of marine bivalves, and also provides a molecular basis for improving the attachment and metamorphosis rate of M. coruscus larvae.