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J. W. Lee, N. Mast, I. Pascual, N. Tserentsoodol, N. B. Javitt, I. A. Pikuleva, N. Gordiyenko, I. R. Rodriguez; Expression and Localization of Cytochrome 46A1 in Monkey Retina. Invest. Ophthalmol. Vis. Sci. 2007;48(13):58.
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© ARVO (1962-2015); The Authors (2016-present)
Cytochrome P450 46A1 (CYP46A1) is known to be a brain-selective sterol hydroxylase that converts cholesterol to 24-hydroxycholesterol for the purpose of maintaining cholesterol homeostasis. The purpose of this study is to demonstrate the expression and localization of CYP46A1 in the monkey retina and to learn about its potential function in the retina.
CYP46A1 protein and mRNA expression were measured in monkey retina and cultured RPE cells by immunoblot analyses and real time RT-PCR. CYP46A1 localization was performed by immunohistochemistry and imaged by confocal microscopy. Subcellular localization was performed by co-transfection of ARPE19 cells with a CYP46A1-GFP construct and pDsRed2 cellular organelle markers. Monkey retina lipids were extracted and analyzed by GC-MS.
Immunoblot analyses detected a significant presence of CYP46A1 enzyme in the monkey retina. Semi-quantitative RT-PCR detected mRNA levels in the retina that were approximately one fifth of those found in the brain. CYP46A1 localized mainly to the photoreceptors outer segments, especially the cones outer segments and synaptic pedicles. Immunoreactivity was also observed in the ganglion cell layer (GCL), RPE and choriocapillaris. CYP46A1-GFP expression in transiently transfected cells localized mainly to the ER but also to the mitochondria. Small amounts of 24-hydroxycholesterol were identified as a TMS derivative by GC-MS in the monkey retina extracts.
CYP46A1 is highly expressed in the photoreceptors, especially the cones outer segments and synaptic pedicles. The localization to cone synaptic pedicles supports the partial mitochondrial localization observed in the transfected ARPE19 cells. The detection of 24-hydroxycholesterol supports its potential function in maintaining cholesterol homeostasis. However, its localization in the photoreceptor outer segments and synaptic pedicles hints at alternative functions.
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