A promoter element with enhancer properties, and the orphan nuclear receptor RORalpha, are required for Purkinje cell-specific expression of a Gi/o modulator.
The promoter and structural portion of the gene, Pcp-2(L7), has frequently been used to target expression of proteins to cerebellar Purkinje cells. In our continuing analysis of the transcription of this gene and how it relates to the G-protein and Ca2+ channel modulatory functions of the encoded protein, we have dissociated the promoter and structural gene and identified cooperative functions. A 0.9 kb fragment of the proximal promoter has positional properties of a classical enhancer, yet its function requires the presence of the structural gene. We demonstrate that RORalpha, the gene product of the mutant mouse locus called staggerer (Rora(sg)), binds to and activates expression through this promoter element using functional assays in vitro and in vivo. The structural gene has a repressive effect on gene expression outside Purkinje cells, and likely participates in the suppression of Pcp-2(L7) gene expression in the many other brain and non-neuronal cell types, besides Purkinje cells, known to express RORalpha. Additional studies in vivo show that while Pcp-2(L7) expression is dependent on RORalpha throughout the cerebellum, this dependence is greatest in the intermediate region between the vermis and far lateral hemispheres. Thus, in addition to its recently indicated role in Ca2+-mediated reciprocal cell-cell signaling in Purkinje cells, RORalpha may also contribute to functional differences in cerebellar subregions.
Serinagaoglu
Yelda
Y
Molecular, Cellular and Developmental Biology Graduate Program, Center for Molecular Neurobiology, The Ohio State University, Columbus, OH 43210, USA.
Zhang
Rui
R
Zhang
Yufang
Y
Zhang
Linda
L
Hartt
Greg
G
Young
Anthony P
AP
Oberdick
John
J
D000818:Animals; D002461:Cell Line, Transformed; D002531:Cerebellum / Q000166:cytology; D047369:Chromatin Immunoprecipitation / Q000379:methods; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D004742:Enhancer Elements, Genetic / Q000502:physiology*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008818:Mice, Neurologic Mutants; D008822:Mice, Transgenic; D009479:Neuropeptides / Q000502:physiology*; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011689:Purkinje Cells / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015534:Trans-Activators / Q000502:physiology*; D014162:Transfection / Q000379:methods
D020662:Guanine Nucleotide Exchange Factors; D009479:Neuropeptides; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; C104971:Pcp2 protein, mouse; C539738:RORA protein, human; D018160:Receptors, Cytoplasmic and Nuclear; C539739:Rora protein, mouse; D015534:Trans-Activators; D019206:GTP-Binding Protein alpha Subunits, Gi-Go