Nonconservative utilization of aldolase A alternative promoters

Recently, analysis of the sequence and expression of the human aldolase A gene revealed the unique arrangement of three tandem promoters and exons preceding a common coding sequence. A muscle-specific promoter (M) and two flanking widely used promoters (N and H) produce mRNA species which, in their mature forms, differ only in the sequence of their 5'-untranslated regions. We have isolated and investigated the expression of a mouse aldolase A gene. This mouse gene represents a functional gene by sequence analysis, recombinational screening, and by transfection into C2C12 cells. Although there is a high degree of sequence similarity between the mouse and the human gene in the region of the alternative first exons, we have been unable to detect a functional utilization of the 5'-most promoter (N) in the mouse. Steady state mRNAs isolated from a variety of adult tissues and cultured cells were analyzed by RNase protection and primer extension to identify first exon utilization. Consistent with previous reports, exon M is found only in skeletal muscle and exon H, the "housekeeping" exon, is utilized in every tissue where aldolase A is expressed. Under identical conditions we fail to see any evidence of the N exon. Therefore, although sequence homology exists between rodents and primates in the N region, the absence of selective pressure to preserve its primate pattern of expression may have resulted in functional promoter extinction.     

Regulation of the osteopontin gene by the orphan nuclear receptor NURR1 in osteoblasts

The orphan nuclear receptor Nurr1 is mainly expressed in the central nervous system but is also detected in certain peripheral tissues such as bone. To elucidate the role of Nurr1 in bone, we examined the ability of Nurr1 to regulate osteopontin (OPN) expression in osteoblastic cell lines. Transfection of Nurr1 in osteoblastic cells increased OPN mRNA expression. A dominant negative Nurr1 variant abolished the ability of PTH to induce OPN expression, suggesting that Nurr1 is involved in mediating the regulation of OPN by PTH. Nurr1 efficiently transactivated a luciferase reporter construct driven by the -857/+191 fragment of the mouse OPN promoter. The activation of the OPN promoter was mediated by the monomeric form of Nurr1, required direct binding of Nurr1 to the OPN promoter, and was dependent on the amino-terminal transactivation function-1. The OPN promoter is also regulated by vitamin D receptor and estrogen-related receptors. We show that Nurr1 and vitamin D activate the OPN promoter in a synergistic fashion, whereas Nurr1-mediated transactivation of the OPN promoter is repressed by estrogen-related receptors. In conclusion, Nurr1 activates the OPN promoter directly in osteoblastic cells, suggesting a role for Nurr1 in the regulation of bone homeostasis.