Abstract
Purpose :
To explore the roles of RP causing proteins in the retinal degeneration mouse model by using proteomic analysis.
Methods :
Sanger sequencing, qPCR and Western blots, combined with morphological and functional assessments (Immunofluorescence, OCT and ERG) illustrated the genotype and phenotype of rd10 mice. Given that retinal structure and functions were partially maintained at 5 weeks, while 8w rd10 mice presented complete loss of retinal function. We applied proteomic analysis to explore the proteins alterations responsible for photoreceptors degeneration in 5w and 8w rd10 mice. In order to assess comparative protein expression, proteins extracted from whole retinas of rd10 and WT were sent to label-free quantitative mass spectrometry. Following proteomic analysis, the significantly reduced expressions of 25 DEPs were further validated by qPCR and Western blots.
Results :
A total of 3737 proteins were identified at the degenerative time points in rd10 mice. 222 and 289 differentially expressed proteins (DEPs) (fold change, FD > 2, p < 0.05) were detected at 5 and 8 weeks, respectively. Based on GO and KEGG pathway analyses, visual perception and phototransduction were severely affected in rd10 mice. The downregulated DEPs were significantly enriched in cilium assembly and protein localization of rd10 mice at 8 weeks. Moreover, 25 decreased DEPs (FD>2, p<0.05) causing autosomal recessive/dominant retinitis pigmentosa were visualized by heatmaps. Protein-protein interaction network represented that 13 DEPs interacted directly with the Pde6b protein. The 25 DEPs responsible for RP were involved with phototransduction, visual perception, response to stimulus, protein localization and cilium assembly pathways. Q-PCR and Western blots verified the changes of DEPs at mRNA and protein levels.
Conclusions :
This study provides an analysis of DEPs causing RP in pde6b mutated mouse model. The results suggested that multiple retinitis pigmentosa associated proteins were involved in retinal degeneration. It revealed the molecular mechanisms underlying early progression and late stage of RP, as well as changes of RP-causing proteins.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.