Expression of Cited2, a Gene Isolated From a Light–induced Retinal Degeneration cDNA Library, in the Degenerating Retina and the Involuting Mammary Gland

Abstract

Abstract: : Purpose: The progression of tissue degeneration leading to controlled cell death is believed to occur by a common genetic mechanism and thus genes expressed in multiple stressed environments may be involved in this common process. To study this hypothesis we utilized Cited2 (Mrg1/p35srj) as a candidate, which was isolated from a light exposed rat retinal cDNA library. Cited2 was also analyzed in the involuting mouse mammary gland (MG), a well studied model of tissue degeneration exhibiting controlled cell death. Methods: A cDNA library was constructed from retinal mRNA isolated from rats exposed to 4 hours of green light. Duplicate plaque lifts were probed with cDNA derived from the retinae of 4 hour light treated rats as well as dark reared rats. Cited2 was identified from the differential screen of genes presumed to be involved in light induced retinal degeneration, LIRD. Cited2 was used to probe Northern blots composed of a LIRD exposure profile and blots composed of the MG developmental profile. In situ analysis was performed on MG involution day 2 sections to determine cellular localization. A polyclonal antibody was created to amino acids 120–135 of the Cited2 protein for western analysis. Results: Expression of Cited2 in the LIRD profile indicates an upregulation during 4 hours of light treatment with peak expression at 8 hours of light exposure. When Cited2 expression was analyzed in the MG, peak expression was witnessed during involution day 2. Western analysis confirmed these results for the MG with expression in early involution. In situ analysis revealed localization of the Cited2 mRNA to the epithelial and potentially myoepithelial cells of the alveoli in the MG. Conclusions: Both degeneration systems analyzed represent phases of apoptotic commitment where the system is deciding whether or not to undergo apoptosis. Both systems involve oxidative stress and tightly regulated tissue degeneration. Thus the expression of Cited2 at similar cellular stress points in both systems alludes to a potential role of this gene in the regulation of the decision to undergo apoptosis. Analysis of genes with common expression profiles in these two tissue degenerating systems may aid in uncovering factors involved in the regulation of tissue degeneration pathways.

Keywords: degenerations/dystrophies • genetics • cell death/apoptosis