Article

Insertion of a Pax6 consensus binding site into the alphaA-crystallin promoter acts as a lens epithelial cell enhancer in transgenic mice.

Center for Molecular and Human Genetics, Columbus Children's Research Institute, Columbus, Ohio 43205, USA.
Investigative Ophthalmology &amp Visual Science (impact factor: 3.6). 07/2004; 45(6):1930-9. pp.1930-9
Source: PubMed

ABSTRACT Although the murine alphaA-crystallin promoter is the most commonly used promoter for achieving transgene expression in the developing lens, this promoter directs transgene expression efficiently only in lens fiber cells. The purpose of the present study was to generate promoters capable of directing transgene expression to the entire lens but not to the corneal epithelium.
Transgenic mice were generated with fragments of the murine alphaA- and alphaB-crystallin promoters, as well as with an alphaA-crystallin promoter engineered with the insertion of a Pax6 consensus binding site driving either human growth hormone (hGH) or Cre recombinase genes. hGH expression was evaluated by in situ hybridization and immunohistochemistry. Cre expression was revealed by x-gal staining after crossing Cre transgenic mice with a Cre reporter strain.
Within the lens, the -214/+38 alphaB-crystallin promoter fragment directed transgene expression in the lens epithelium, but not in fiber cells. The native -282/+43 alphaA-crystallin promoter drove transgene expression in the lens fiber cells of several independent lines of transgenic mice, but none of these mice demonstrated significant transgene expression in the lens epithelium. In contrast, the insertion of a 32-bp sequence containing a Pax6 consensus binding site into the -282/+43 alphaA-crystallin promoter reproducibly led to transgene expression in the lens epithelium as well as the lens fiber cells.
The inclusion of a Pax6 consensus binding site within the -282/+43 alphaA-crystallin promoter enhances the ability of this promoter to drive transgene expression in the lens epithelium.

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Keywords

-214/+38 alphaB-crystallin promoter fragment
 
-282/+43 alphaA-crystallin promoter enhances
 
32-bp sequence
 
alphaB-crystallin promoters
 
corneal epithelium
 
Cre expression
 
Cre reporter strain
 
Cre transgenic mice
 
developing lens
 
drive transgene expression
 
entire lens
 
hGH expression
 
independent lines
 
lens epithelium
 
lens fiber cells
 
murine alphaA-
 
murine alphaA-crystallin promoter
 
significant transgene expression
 
situ hybridization
 
transgene expression