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RE Hausman, RS Mukherjee; Embryonic Expression of the Chicken Choline Acetyltransferase Gene Complex . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2446.
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Purpose:Choline acetyltransferase (ChAT) is the enzyme that synthesizes acetylcholine (ACh) and the vesicular acetylcholine transporter (VAChT) is the pump that packages ACh into vesicles. ACh is a morphogen early in retina development and a neurotransmitter in the adult. It is subject to, at least, two forms of control from outside the cell, by growth factors and via cell contacts. Previous work shows that this control is both translational and transcriptional. To understand the developmental control of this gene complex, we have characterized the expression of both messages during chick embryo retina development. Methods:Randomly-labeled probes generated from a rat ChAT cDNA were used to screen a chick brain cDNA library. From the chicken ChAT cDNA isolated, randomly-labeled probes were generated for genomic library screening. Additional primers were designed to sequence both chicken ChAT and the nested vesicular ACh transporter (VAChT) and for use in northern analysis. Results:Approximately 90% of the expected VAChT-ChAT complex gene has been sequenced and the results show strong sequence similarity with the known mammalian gene complexes. In both embryonic retina and forebrain we detected a single major message for each of the proteins and these appeared to be expressed in post hatch chicken cerebrum. RT-PCR shows that the ChAT message contains the non-coding M exon. During early embryonic development the increase in mRNA level for both proteins tracked in increase in protein level in both retina and forebrain. However, by about E11 there was a loss of correlation. Conclusion:The VAChT-ChAT gene complex is strongly conserved when compared with that of mammals. By northern blotting, there is only one major ChAT and VAChT message respectively in chick brain and retina. The ChAT message is developmentally regulated but after the initial appearance of expression loses its parallel to protein level suggesting that the major control during early embryonic development may be at the mRNA level but later in embryogenesis there must be additional post-transcriptional regulation.
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