Characterization and genomic structure of Dnah9, and its roles in nodal signaling pathways in the Japanese flounder (Paralichthys olivaceus).

The nodal signaling pathway has been shown to play crucial roles in inducing and patterning the mesoderm and endoderm, as well as in regulating neurogenesis and left-right axis asymmetry. Here, we present the first complete cDNA and genomic sequences as well as the promoter predication of the Dnah9 gene in the Japanese flounder. The 15,558-bp-long cDNA is divided into 96 exons and spread over 138 kb of genomic DNA. Protein sequence comparison showed that it shares higher identity with other vertebrate orthologs, with an ATP binding dynein motor, AAA domain and microtubule binding stalk of dynein motor. Dnah9 exhibited maternal and ubiquitous expression in all cells of the early development stages, but became concentrated in the head at 1 DAH, as identified by qRT-PCR and in situ hybridization methods. Furthermore, after nodal signaling was inhibited, the level of Southpaw did not change significantly at early development stage (50 % epiboly) but increased significantly at late stages (27-somite stages and 1 DAH), as well as the expression of Lefty, an inhibitor of nodal signaling, increased continuously. On the other hand, the expression level of Dnah9 decreased. The transcription factor binding site of FAST-1 (SMAD interacting protein) was identified in the transcription region of Dnah9 by the promoter analysis, which might format the complexes of SMADs, FAST-1 and the transcription region of Dnah9 served as a bridge of Dnah9 and nodal signaling. All evidences indicated that Dnah9 might be downstream of nodal during the early development stages, and an indirect function through SMADs for nodal signaling pathway.