DISSECTING THE TRANSCRIPTIONAL CODE OF THE DOPAMINE D2 RECEPTOR GENE

LORENZO RAMIRO; RUBINSTEIN MARCELO

Rosario

Congreso; 50th. Annual Meeting Argentine Society for Biochemistry and Molecular Biology; 2014

Sociedad Argentina de Investigación Bioquímica y Biología Molecular

The dopamine D2 receptor (D2R) plays a major role in the central control of locomotor, appetitive, emotional and cognitive functions. Despite its significance, the molecular mechanisms that control the expression of the D2R gene (Drd2) in different brain areas remain unknown. BAC transgenic mice carrying a genomic fragment of 220 kb containing Drd2 and EGFP coding sequences (BAC BX37), express the reporter protein in most brain areas where the D2R is normally found. Using BAC engineering we shortened this recombinant BAC to 60 kb. Transgenic mice carrying the 60 kb BAC showed an EGFP expression pattern that was indistinguishable from BX37 mice. To isolate regulatory elements capable of directing the expression into diverse subpopulations of neurons expressing Drd2 we further dissected DNA regions based on sequence conservation criteria. Putative regulatory regions were subcloned upstream of a minimal promoter followed by coding sequences of a red fluorescent protein gene. Analysis of transgenic mouse lines carrying different conserved sequences showed expression of the reporter gene in distinct regions including the septum, endopiriform nucleus, striatum, olfactory tubercle and midbrain. Thus, this methodology seems to be effective for the molecular dissection of transcriptional enhancers of a complex gene.