Molecular, functional, and gene expression analysis of zebrafish Ror1 receptor
Ror family of receptor tyrosine kinases ROR1 and ROR2 plays crucial roles in animal development by regulating cell proliferation, differentiation, and migration, as well as survival and death by acting as a receptor or co-receptor for Wnt5a and mediating Wnt5a-induced activation. Compared with our extensive understanding of ROR2, our knowledge of ROR1 is limited. In this study, we characterized the zebrafish ror1 gene and determined its temporal and spatial expression and biological activity. Sequence comparison and phylogenetic analyses indicate that its protein structure is similar to its mammalian orthologs. During embryogenesis, the ror1 mRNA levels were relatively low or undetectable at 6 and 9 h postfertilization. In adult fish, ror1 mRNA was most abundantly expressed in the ovary and testis. The levels of ror1 mRNA in non-reproductive system tissues were very low or barely detectable. Spatiotemporal distribution of ror1 and its ligand wnt5a in the ovary was then investigated. Reverse transcription PCR on isolated follicle layers and denuded oocytes demonstrated that both wnt5a and ror1 were exclusively expressed in the oocyte but not in the follicle layers. During oogenesis, the ror1 mRNA levels were relatively low from I to IV stage oocytes and increased dramatically at V stage oocyte. Unlike ror1, the wnt5a mRNA levels were increased gradually from I to V stage oocyte. When Ror1 was co-transfected with Wnt5a and Wnt3a in HEK293T cells, the Wnt3a-induced Wnt reporter activity was inhibited by Ror1 in a dose-dependent manner. Taken together, these results provide new information about the structural and functional conservation, spatial and temporal expression, and biological activity of Ror1 in a fish model organism.
KeywordsRor1 Zebrafish Gene expression
This work was supported by grants from the National Science Foundation of China (Grant # 31001099 and 30972238) and partially supported by the Fundamental Research Funds for the Central Universities (CZZ17004).
- Abedini A, Zamberlam G, Lapointe E, Tourigny C, Boyer A, Paquet M, Hayashi K, Honda H, Kikuchi A, Price C, Boerboom D (2016) WNT5a is required for normal ovarian follicle development and antagonizes gonadotropin responsiveness in granulosa cells by suppressing canonical WNT signaling. FASEB J 30(4):1534–1547CrossRefGoogle Scholar
- Endo M, Minami Y (2018) Diverse roles for the Ror-family receptor tyrosine kinases in neurons and glial cells during development and repair of the nervous system. In: Dev Dyn, vol 247, pp 24–32Google Scholar
- Fukuda T, Chen L, Endo T, Tang L, Lu D, Castro JE, Widhopf GF 2nd, Rassenti LZ, Cantwell MJ, Prussak CE, Carson DA, Kipps TJ (2008) Antisera induced by infusions of autologous Ad-CD154-leukemia B cells identify ROR1 as an oncofetal antigen and receptor for Wnt5a. Proc Natl Acad Sci U S A 105(8):3047–3052CrossRefGoogle Scholar
- Nomi M, Oishi I, Kani S, Suzuki H, Matsuda T, Yoda A, Kitamura M, Itoh K, Takeuchi S, Takeda K, Akira S, Ikeya M, Takada S, Minami Y (2001) Loss of mRor1 enhances the heart and skeletal abnormalities in mRor2-deficient mice: redundant and pleiotropic functions of mRor1 and mRor2 receptor tyrosine kinases. Mol Cell Biol 21(24):8329–8335CrossRefGoogle Scholar