Abstract
Purpose: :
To investigate the effect of Bves (zBVES) in the eye development and find the potential mechanism which Bves influence through.
Methods: :
Tol-2 transposon system was established to produce a transgenic zBves promoter-EGFP zebrafish line, and then the fish was observed by a confocol microscope to investigate the expression pattern of Bves in eye. To examine its function, knockdown of zBves that created by injecting embryos with MOs was applied. Transmission electron microscope (TEM) technique and immunostaing with antibodies- zn8, anti-corbonic anhydrase (CA), anti-tyrosine hydroxylase (TH), and Zpr-1-were adopted to identify the defeats of zBves knockdown embryo in eyes. Further, we used in situ hybridization to study the temporal and spatial expression of various genes-Otx2, Rx1, Six3b, Pax6a, Pax2a, and Vsx2- in different eye developing stages after zBves knockdown. To confirm the relevance between sonic hedgehog (Shh), wnt, and zBves, LiCl treatment (GSK3 inhibitor) and co-injecting Shh or Wnt2 messenger RNAs were performed with knockdown embryos.
Results: :
The transgenic zebrafish line revealed that zBves was expressed in corneas, lenses, and retinas of zebrafish eyes. After knockdown of zBves, the eye could not be formed maturely and was resulted in a microphthalmos which was shown that photoreceptor layer was not formed in the immunostaining and TEM. Furthermore, Pax6a disappeared and Pax2a changed its expression pattern at 10-12hpf, early eye field formation, and the decreased expressions of Pax6a, Pax2a, and Vxs2 were also found at 24hpf, neural retina formation. LiCl treatment at 10-12hpf could rescue the microphthalmos phenomenon in zBves knockdown embryos.
Conclusions: :
zBves could influence eye development via Pax6a and Pax2a under the effects of Wnt or Shh.
Keywords: photoreceptors • transcription factors