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K.M. Veldhoen, W.T. Allison, C.W. Hawryshyn; Proteomic Analysis of Retinal Development Using ICAT . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4540.
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Purpose: We are using thyroid hormone treatment to mimic natural metamorphosis of the rainbow trout. This approach provides us with a powerful model to study photoreceptor cell death and regeneration in the retina. Previous data indicate that thyroid hormone induces apoptosis of ultraviolet-sensitive cones, generation of rod photoreceptors and stretching/compaction of the retinal mosaic. We are using a proteomic approach to identify and characterize the components involved in these cellular events. Methods: Retinal homegenates were prepared from control and thyroid hormone-treated fish. Peptides from control and treated homogenates were differentially labelled using isotope-code affinity tags (ICAT) and analyzed using capillary liquid chromatography-electrospray ionization-tandem mass spectrometry (capLC-ESI-MS/MS). This method identifies proteins and determines their relative abundance between samples. Results: Thyroid hormone affected the relative abundance of photoreceptor-specific proteins. Examples include an observed increase in arrestins and rod opsin. We also observed an increase in proteins associated with differentiation/regeneration of retinal neurons. In addition, there were 17 unidentified peptides (not matched to proteomic databases) that increased more than five-fold in abundance, and 21 unidentified peptides that decreased more than five-fold in abundance following thyroid hormone treatment. Manual sequencing of the MS/MS spectra of these peptides will allow further characterization of the role of specific proteins in retinal development. Conclusions: We have shown that there are differences in the proteomes of control and thyroid hormone-treated retinae. These differences have identified protein targets for further investigation. ICAT and capLC-ESI-MS/MS serve as an effective compliment to other molecular biological approaches that help to elucidate the mechanisms of retinal development. Supported by a Fellowship from the Alzheimer Society of Canada and the CIHR Institute of Aging (WTA) & operating grants from NSERC (CWH).
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