This study has attempted to identify regions of DNA within the VIP gene that may be responsible for spontaneous expression of VIP. Using an electrophoretic mobility shift assay, this study has shown that the rat VIP CRE is capable of binding c-Jun in a heterodimer with c-Fos. To determine the importance of these proteins in binding to the VIP CRE, an attempt was made to compete them off the endogenous rat VIP CRE. DRG neurons were transfected with constructs containing copies of this CRE ligated into the plasmid pUC18. Quantitative analysis of the effects of transfection on endogenous VIP immunoreactivity, showed that the CRE containing construct caused a selective reduction in VIP expression. Proteins binding to the CRE are therefore important for spontaneous VIP expression. To determine if the rat VIP CRE is all that is necessary for spontaneous VIP expression, it was analysed, using reporter constructs, for its ability to mediate patterns of gene expression analogous to those seen for endogenous VIP. This study has shown that the CRE is capable of increasing gene transcription from a heterologous c-fos promoter in neonatal, but not adult rat DRG neurons, in response to stimuli that raise cAMP and intracellular calcium and as such may be responsible for the synergistic increase in VIP expression that occurs in neonatal rat DRG neurons in response to these same stimuli. This study suggests that the spontaneous expression of rat VIP is dependent on protein complexes binding to the CRE, and that these complexes probably contain c-Jun. Although the CRE alone is capable of mediating the response of VIP to cAMP and calcium, and may mediate developmental differences in VIP expression, sequences are required in combination with the CRE for this spontaneous expression.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:649587 |
| Date | January 1996 |
| Creators | Dobson, Stephen P. |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/21200 |
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