Enhanced retinal ganglion cell differentiation by ath5 and NSCL1 coexpression.
OBJECTIVE
The molecular mechanism underlying retinal ganglion cell (RGC) differentiation is not fully understood. In this study, the role of the basic helix-loop-helix (bHLH) genes ath5 and NSCL1 in RGC differentiation was examined, by testing whether their coexpression would promote RGC differentiation to a greater extent than either gene alone.


METHODS
The replication-competent avian RCAS retrovirus was used to coexpress ath5 and NSCL1 through an internal ribosomal entry site. The effect of the coexpression on RGC differentiation was assayed in vivo in the developing chick retina and in vitro in RPE cell cultures derived from day 6 chick embryos.


RESULTS
Coexpression of ath5 and NSCL1 in RPE cells cultured in the presence of bFGF promoted RPE transdifferentiation toward RGCs, and the degree of transdifferentiation was much higher than with either gene alone. Cells expressing RGC markers, including RA4, calretinin, and two neurofilament-associated proteins, displayed processes that were remarkably long and thin and often had numerous branches, characteristics of long-projecting RGCs. In the developing chick retina, retroviral expression of NSCL1 resulted in a moderate increase in the number of RGCs, results similar to retroviral expression of ath5. Coexpression of ath5 and NSCL1 yielded increases in RGCs greater than the sum of their increases when expressed separately.


CONCLUSIONS
Both in vitro and in vivo data indicate that the combination of ath5 and NSCL1 promotes RGC differentiation to a greater degree than either gene alone, suggesting a synergism between ath5 and NSCL1 in advancing RGC development.
Department of Ophthalmology, University of Alabama at Birmingham, Birmingham, Alabama 35233, USA.
Wenlian
W
Xie
Run-Tao
RT
Yan
Wenxin
W
Ma
Shu-Zhen
SZ
Wang
D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D002642:Chick Embryo; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D012165:Retinal Ganglion Cells / Q000166:cytology* / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D014157:Transcription Factors