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Ameair Abu Irqeba, Judy M Ogilvie; The Precise Trafficking of Prenylated Rab Acceptor 1 in Photoreceptors is Dependent Upon Both of its Cytoplasmic Domains. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5515. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The rd1 mouse is a well-studied model of Retinitis Pigmentosa (RP), a disease in which photoreceptors degenerate. We conducted a microarray analysis comparing rd1 with WT retina and have previously demonstrated that prenylated rab acceptor 1 (PRA1), a four pass transmembrane protein, is consistently down-regulated at both the mRNA and protein level before postnatal day ten. We also observed that at the onset of degeneration, the golgi apparatus is fragmented and PRA1 is mis-localized within the inner segment. The role that PRA1 plays in photoreceptors is unknown, but previous reports have alluded to its involvement in retrograde trafficking of Rab GTPases. Here, we examine the importance of its cytoplasmic domains for proper localization within photoreceptors.
To gain insight into the mechanism through which PRA1 is trafficked within photoreceptors, we truncated the protein. Primers were designed to amplify PRA1 without its N-terminal cytoplasmic domain, without its C-terminal cytoplasmic domain, or without either of these amino acid sequences. PCR constructs were sub-cloned into a vector where they were tagged with EGFP (under the control of the CAG promoter). They were then injected subretinally into wild type retina and electroplated at P0.
We observed that the loss of either the N- or C-terminal cytoplasmic domain did not lead to the secretion of PRA1 into the outer segment. Loss of both the N- and C-terminal domains together did lead to the movement of PRA1 into the outer segment. In photoreceptors, golgi retention seemed to rely on either the N- or C-terminus, loss of both leads to an expedient exit from the golgi. Loss of transmembrane regions three and four leads to a localization phenotype similar to that of WT PRA1.
We show here that both of PRA1’s cytoplasmic domains are important for its proper trafficking within photoreceptors, although the C-terminus seems to be essential for ER exit. Normal golgi localization correlates with proper overall trafficking within photoreceptors. This supports our hypothesis that an important interaction occurs within the golgi, possibly utilizing these cytoplasmic domains, that facilitates PRA1 trafficking within the inner segment and to the ribbon synapse.
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