IRE1-XBP1 pathway in mutant T4R <i>RHO</i> and <i>WT</i> canine retinas 6 hours after light exposure.

<p><b>(A)</b> RT-PCR analysis of <i>XBP1</i> splicing in light exposed (E) compared to shielded (S) T4R <i>RHO</i> and <i>WT</i> retinas. RT-PCR of canine <i>XBP1</i> generated a 289 bp fragment, which represents the unspliced form of canine <i>XBP1</i>. The 263 bp fragment, which represents the spliced form of canine <i>XBP1</i> was not observed except in the tunicamycin treated normal canine fibloblasts (NCF). A retina from a wild-type dog kept under standard ambient kennel illumination (K) was used as a control of basal <i>XBP1</i> expression and splicing. <b>(B)</b> Differential expression of genes <i>XBP1 and ASK1</i> in the retinas of three RHO ^T4R/T4R mutant dogs following light exposure. Three different sets of primers were used to specifically amplify the unspliced (u), spliced (s) and both (total) <i>XBP1</i> transcripts. Displayed are the mean fold change (FC) difference compared to the contralateral shielded retinas. Error bars represent the FC range (FC min to FC max). <b>(C)</b> Immunoblots showing the protein levels of total and phosphorylated forms of XBP1 in light exposed (E) compared to shielded (S) retinas of mutant (<i>RHO</i>^T4R/T4R, and <i>RHO</i>^T4R/+), and <i>WT</i> (+/+) dogs. A single retina from a wild-type dog kept under standard ambient kennel illumination (K) was included as a control of basal levels of XBP1.