Expression And Target Predictions Of Let-7d During Metamorphosis In Japanese Flounder (Paralichthys Olivaceus)

The Japanese flounder is one of the most commercially important Pleuronectid flatfish. There is a metamorphosis from bilaterally symmetric larvae to asymmetric juveniles in its early life history. The right eye moves to the left side of the body, and the two eyes become located only on one side, which is the most typical left-right asymmetric phenomenon seen in vertebrate. It involves to many modification of morphological structure, physiological function, biochemical elements and behavior style.MicroRNA (miRNA) is the smallest, functional, noncoding RNAs of plants and animals. Mature miRNAs are ~22 nt in length and are cleaved from the ~75 nt partially duplexed precursor by the RNase III Dicer. The first to be discovered were the lin-4 and let-7, which are components of the gene regulatory network that controls the timing of C.elegans from larval development. MiRNA have been found in C.elegans, Drosophila, plants, mice and humans, suggesting an ancient and widespread role for these non-coding RNAs. The high sequence conservation of many miRNAs across metazoan species suggests strong evolutionary pressure and participation in essential biologic processes. Indeed, miRNA have been shown to play fundamental roles in diverse biological and pathological processes, including cell proliferation, differentiation, apoptosis, and carcinogenesis. In contrast to plant miRNAs, which usually bind nearly perfectly to their targets, animals miRNA–target interactions occur through semicomplementary base pairing, usually within the 3'UTRs of the target genes. In animals, miRNAs usually inhibit gene expression through partially complementary elements in the 3'-UTRs of their target mRNAs. MiRNA let-7 is a critical regulator of developmental timing events at the larval-to-adult transition, especially in Drosophila melanogaster metamorphosis. MiRNAs in Paralichthys olivaceus were identified by Solexa sequencing. Ten miRNAs belonged to the let-7 family (from let-7a to let-7j). The copy numbers of let-7d were significantly higher than the other members of the let-7 family. Quantitative analysis revealed that the expression levels of let-7d in the normal control group and the thyroid hormones (THs) treatment group had a common characteristic: expression gradually increased to a peak level, and then decreased to a low level. Let-7d levels in the THs treatment group were 50% less than those of the normal control group at each stage of development (P < 0.05). let-7d was detected in all tissues examined, whereas at relatively higher levels in gill, heart and brain. Paralichthys olivaceus Precursor-let-7d (Pre-let-7d) was cloning from genoe DNA. Alignment analysis showed Pre-let-7d was familiar with other organism Pre-let-7d. Data of target predictions indicated that the 3'-untranslated regions (3'-UTRs) of 30 registered mRNAs existed target sites matched with let-7d. The results of the present study suggested that let-7d played an essential role in regulating the developmental timing and remodeling the body structures of Paralichthys olivaceus during metamorphosis. The highly expressed let-7d could interact with a number of genes and regulate these genes by acting on 3'-UTRs of genes, participating in the regulation of Paralichthys olivaceus metamorphosis.