The human cGMP-PDE beta-subunit promoter region directs expression of the gene to mouse photoreceptors.
OBJECTIVE
We previously demonstrated that 350 bp of the human rod cGMP phosphodiesterase beta-subunit (beta-PDE) gene promoter are sufficient to direct high levels of gene expression in human Y-79 retinoblastoma cells in vitro. In this study the cell specificity and expression pattern conferred by the short beta-PDE 5' flanking sequence in vivo were examined.


METHODS
A construct containing the bacterial LacZ gene driven by a fragment of the beta-PDE 5' flanking region (-297 to +53) was used to generate transgenic mice. Gene expression was analyzed by measuring beta-galactosidase activity in tissue homogenates or visualizing enzymatic activity or protein production at a cellular level by in situ histochemistry or immunocytochemistry.


RESULTS
Three independently derived transgenic lines were generated carrying the -297 to +53 beta-PDE 5' flanking region fragment. Within the retina, the reporter gene was specifically expressed in photoreceptors, consistent with the localization of endogenous beta-PDE. Significant expression of LacZ was not observed in other ocular or peripheral tissues.


CONCLUSIONS
Photoreceptor-specific reporter gene expression is driven in vivo by a 350-bp segment of the beta-PDE 5' flanking sequence. This study demonstrates the utility of the human beta-PDE promoter for directing the expression of foreign genes to photoreceptors and suggests that the -297 to +53 beta-PDE 5' flanking region fragment may have important implications for therapeutic gene delivery to the visual cells.
Ogueta
S B
SB
Department of Ophthalmology and Jules Stein Eye Institute, UCLA School of Medicine, Los Angeles, California, USA.
Di Polo
A
A
Flannery
J G
JG
Yamashita
C K
CK
Farber
D B
DB
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000096:biosynthesis / Q000235:genetics*; D000818:Animals; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D017930:Genes, Reporter / Q000235:genetics; D007763:Lac Operon / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D010727:Phosphoric Diester Hydrolases; D020419:Photoreceptor Cells, Vertebrate / Q000201:enzymology*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014018:Tissue Distribution; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D010727:Phosphoric Diester Hydrolases; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; C517431:PDE6B protein, human; C517438:Pde6b protein, mouse; D001616:beta-Galactosidase