Figure 11.

Differential regulation of Utrn‐A and Utrn‐B promoters in the central nervous system. Expression pattern of Utrn‐A and Utrn‐B transcripts (A,B) and endogenous promoter activity (C,D) was studied in primary cultures of neurons and astroglia isolated from mdx mouse brain, C2C12 and bEnd.3 cell lines. A,B. Utrn‐A transcript level was significantly higher (n = 6; *P < 0.05) in astrocytes and neurons compared with C2C12, whereas in bEnd.3 cells Utrn‐A mRNA was significantly lower (n = 6; *P < 0.05). Utrn‐B mRNA expression was significantly (n = 6; **P < 0.001) higher in bEnd.3 cells, whereas neurons and astrocytes showed significantly (n = 6; **P < 0.001) lower amount of Utrn‐B mRNA compared with C2C12 cells. Mean ± SD are shown. Statistical analysis was conducted using Student's t‐tests. C,D. Differential expression of endogenous Utrn‐A and Utrn‐B promoter activity. Primary neurons, astrocytes, C2C12 cell and bEnd.3 cell cultures were transfected with utrophin‐A (mUtrn‐A‐luc) and/or Utrn‐B (mUtrn‐B‐luc) reporter constructs, along with transfection control pRL‐TK. Promoter activity was assayed 24 h after trasnfection. Endogenous Utrn‐A promoter activity was the greatest in neurons and significantly higher than astrocytes compared with C2C12 cells or bEnd.3 cells. Endogenous Utrn‐B promoter activity was highest in bEnd.3 cells, and significantly high in neurons and astrocytes compared with C2C12 cells. Utrn‐A or Utrn‐B‐derived firefly luciferase activities were normalized to pRL‐TK‐derived renilla luciferase activity (internal control) and expressed as 100% to C2C12 cells. Normalized luciferase values are mean values of 6 wells in three separate experiments. Mean ± SD is shown. Statistical analysis was conducted using Student's t‐tests. Schematic representations of mouse utrophin promoters are shown. Schematic representation of regions of mouse Utrn‐A and Utrn‐B promoter cloned to generate luciferase reporter constructs is given further.