DOI: 10.5061/dryad.c2fqz61pc
title: Retinal proteome profiling of inherited retinal degeneration across three different mouse models suggests common drug targets in retinitis pigmentosa

description: Inherited retinal degenerations (IRDs) are a leading cause of blindness
 among the population of young people in the developed world. Approximately
 half of IRDs initially manifest as a gradual loss of night vision and
 visual fields, characteristic of retinitis pigmentosa (RP). Due to
 challenges in genetic testing and the large heterogeneity of mutations
 underlying RP, targeted gene therapies are an impractical large-scale
 solution in the foreseeable future. For this reason, identifying key
 pathophysiological pathways in IRDs that could be targets for
 mutation-agnostic and disease-modifying therapies (DMTs) is warranted. In
 this study, we investigated the retinal proteome of three distinct IRD
 mouse models, in comparison to sex- and age-matched wild-type mice.
 Specifically, we used the Pde6βRd10 (rd10) and
 RhoP23H/WT (P23H) mouse models of autosomal recessive and
 autosomal dominant RP, respectively, as well as the Rpe65^-/-^ mouse model
 of Leber's congenital amaurosis type 2 (LCA2). The mice were housed
 at two distinct institutions and analyzed using LC-MS in three separate
 facilities/instruments following data-dependent and data-independent
 acquisition modes. This cross-institutional and multi-methodological
 approach signifies the reliability and reproducibility of the results. The
 large-scale profiling of the retinal proteome, coupled with in vivo
 electroretinography recordings, provided us with a reliable basis for
 comparing the disease phenotypes and severity. Despite evident
 inflammation, cellular stress, and downscaled phototransduction observed
 consistently across all three models, the underlying pathologies of RP and
 LCA2 displayed many differences, sharing only four general KEGG pathways.
 The opposite is true for the two RP models in which we identify remarkable
 convergence in proteomic phenotype, even though the mechanism of primary
 rod death in rd10 and P23H mice is different. Our data highlights the cAMP
 and cGMP second-messenger signaling pathways as potential targets for
 therapeutic intervention. The proteomic data are curated and made publicly
 available, facilitating the discovery of universal therapeutic targets for
 RP.
description: Animal Models and Study Design We used three
 different mouse models of inherited retinal degenerative diseases (IRD) to
 discover retinal proteomic changes that are common to retinal degeneration
 (RD). The models used in this study were
 B6.CXB1-<em>Pde6b</em><sup>rd10</sup>/J (RRID:
 IMSR_JAX:004,297, referred to as rd10),
 B6.129S6(Cg)-<em>Rho</em><sup>tm1.1Kpal</sup>/J
 (RRID: IMSR_JAX:017,628, referred to as P23H), mouse models of recessive
 and autosomal dominant retinitis pigmentosa (RP), respectively (18, 19), and B6.129-<em>Rpe65</em><sup>tm1Tmr</sup>/J (RRID: IMSR_JAX:035,329, referred to as <em>Rpe65</em><sup>−/−</sup>) model of Leber congenital amaurosis type 2 (LCA2) which was a kind gift from Dr Michael Redmond (National Institutes of Health) (20). The rd10 colony was kept as a homozygote. Age- and sex-matched C57BL/6J mice (RRID: IMSR_JAX:000,664) were used as controls. P23H heterozygote mice were bred with C57BL/6J mice, yielding P23H heterozygote and wild-type (WT) littermates. To get <em>Rpe65</em><sup>−/−</sup> and their WT littermate mice, we bred heterozygote <em>Rpe65</em><sup>+/−</sup> mice together. Only WT and homozygote offspring were used in this study. Several cohorts of mice were raised, and their retinal samples were collected at two different institutions: the University of California, Irvine (UCI) and the University of Eastern Finland (UEF) over the years 2019 to 2021 and 2023, respectively (Table 1). Mice were given water and standard feed <em>ad libitum</em> at both institutions. Retinal proteome analysis was conducted using liquid chromatography-tandem mass spectrometry (LC-MS/MS), employing data-dependent acquisition (DDA) or data-independent acquisition (DIA) modes. Mice reared P0-P29 in DR and in CLR (vivarium) P29-P38.
description: # Retinal proteome profiling of inherited retinal degeneration across
 three different mouse models suggests common drug targets in retinitis
 pigmentosa Dataset DOI: [10.5061/dryad.c2fqz61pc](10.5061/dryad.c2fqz61pc)
 ## Description of the data and file structure ### Files and variables This
 dataset is showing all proteins identified in indicated group of samples,
 combined by genotype/group. **Files (data generated at UCI, US):** *
 protDDA_Rd10_DR.csv * protDDA_Rd10_CLR.csv * protDDA_P23H.csv *
 protDDA_RPE65_KO.csv **Variables:** * P23H_het_M - heterozygous mice for
 P23H mutation in Rhodopsin, Male * P23H_het_F - heterozygous mice for P23H
 mutation in Rhodopsin, Female * Rd10_M - A mouse genotype with a point
 mutation in the Pde6b gene, leading to progressive photoreceptor
 (especially rod) degeneration, used as a model for retinitis pigmentosa,
 Male * Rd10_F - A mouse genotype with a point mutation in the Pde6b gene,
 leading to progressive photoreceptor (especially rod) degeneration, used
 as a model for retinitis pigmentosa, Female * Rd10_M_DR - A mouse genotype
 with a point mutation in the Pde6b gene, leading to progressive
 photoreceptor (especially rod) degeneration, used as a model for retinitis
 pigmentosa, Male, Dark Reared * Rd10_F_DR - A mouse genotype with a point
 mutation in the Pde6b gene, leading to progressive photoreceptor
 (especially rod) degeneration, used as a model for retinitis pigmentosa,
 Female, Dark Reared * RPE65_M - Mice lack functional RPE65, leading to
 impaired visual cycle, absence of 11-cis-retinal, and progressive retinal
 degeneration, Male * RPE65_F - Mice lack functional RPE65, leading to
 impaired visual cycle, absence of 11-cis-retinal, and progressive retinal
 degeneration, Female * WT_M - wild type mice, Male * WT_F - wild type
 mice, Female * **Razor...unique.peptides** Number of peptides used for
 quantification: unique to the protein or shared with a minimal number of
 proteins ("razor" peptides). * **Unique.peptides** Number of
 peptides that are unique to a single protein group (not shared with
 others). * **Sequence.coverage....** Percentage of the protein sequence
 covered by all identified peptides. *
 **Unique...razor.sequence.coverage....** Sequence coverage based only on
 unique and razor peptides. * **Unique.sequence.coverage....** Sequence
 coverage based only on unique peptides. * **Mol..weight..kDa.** Molecular
 weight of the protein in kilodaltons (kDa). * **Q.value** Adjusted p-value
 (false discovery rate); measures the confidence of protein/peptide
 identification. * **Score** A scoring value indicating the quality or
 confidence of peptide/protein identification (algorithm-dependent). *
 **Intensity** Total signal intensity of the peptides matched to a protein;
 often used for quantification. * **iBAQ.peptides** Number of theoretically
 observable peptides used in iBAQ (intensity-Based Absolute Quantification)
 calculation. * **MS.MS.count** Number of tandem MS (MS/MS) spectra that
 have been assigned to peptides from the protein — indicates identification
 confidence. * **Protein.IDs** List of all protein identifiers (e.g.,
 UniProt IDs) associated with a protein group detected in the experiment. *
 **Majority.protein.IDs** The main (majority) protein ID(s) among the group
 — used as a representative identifier for the protein group. *
 **Protein.names** Common or descriptive names of the proteins (from
 databases like UniProt). * **Gene.names** Standard gene symbols
 corresponding to the proteins (e.g., ACTB, GAPDH). * **WT_vs_P23H_logFC**
 Log₂ fold change of protein abundance between two conditions — here,
 wild-type (WT) vs. P23H (likely a disease model or mutant). Positive =
 upregulated in P23H; negative = downregulated. * **id** Unique identifier
 for each row (protein group) in the dataset, usually automatically
 generated by the analysis software. **Files:** **data generated at UEF,
 Finland** * protDIA_Rd10_DR.csv * protDIA_P23H.csv Variables * P23H_het_M
 - heterozygous mice for P23H mutation in Rhodopsin, Male * P23H_het_F -
 heterozygous mice for P23H mutation in Rhodopsin, Female * Rd10_M_DR - A
 mouse genotype with a point mutation in the Pde6b gene, leading to
 progressive photoreceptor (especially rod) degeneration, used as a model
 for retinitis pigmentosa, Male, Dark Reared * Rd10_F_DR - A mouse genotype
 with a point mutation in the Pde6b gene, leading to progressive
 photoreceptor (especially rod) degeneration, used as a model for retinitis
 pigmentosa, Female, Dark Reared * WT_M - wild type mice, Male * WT_F -
 wild type mice, Female **Statistical comparison columns:**  *
 **WT_vs_(GENOTYPE)_AveExpr** Average expression level of the protein
 across all samples (usually log₂-transformed intensity). *
 **WT_vs_(GENOTYPE)_t** t-statistic from differential expression analysis
 between WT and P23H groups — reflects the strength and direction of
 change. * **WT_vs_(GENOTYPE)_P.Value** Raw p-value indicating the
 probability that the observed difference is due to chance. *
 **WT_vs_(GENOTYPE)_adj.P.Val** Adjusted p-value (e.g., FDR) correcting for
 multiple testing — used to assess significance more reliably. *
 **WT_vs_(GENOTYPE)_B** B-statistic or log-odds that a protein is
 differentially expressed (higher = more likely DE); used in limma-based
 analyses. * **WT_vs_(GENOTYPE)_-log(P.Value)** Negative log₁₀ of the
 p-value — larger values indicate stronger statistical significance (e.g.,
 for volcano plots). * **WT_vs_(GENOTYPE)_-log(adj.P.Val)** Negative log₁₀
 of the adjusted p-value — used similarly to emphasize significance after
 correction. **Protein information columns:** * **Protein.Group**
 Identifier for a group of proteins that cannot be distinguished based on
 peptides (typically grouped by shared peptides). * **Protein.Ids** UniProt
 or database accession numbers for proteins in the group. *
 **Protein.Names** Common or descriptive names of the proteins in the
 group. * **Genes** Gene symbols associated with the proteins (e.g., RHO,
 GAPDH). * **WT_vs_(GENOTYPE)_logFC** Log₂ fold change in protein abundance
 between **(GENOTYPE)** and WT. Positive = higher in **(GENOTYPE)**
 Negative = lower in **(GENOTYPE)** * **First.Protein.Description**
 Detailed description of the first (or main) protein in the group —
 includes functional annotation, organism, etc. #### This dataset is
 showing all pathways identified in indicated group of samples, combined by
 genotype/group. **Files** * pathDIA_P23H.csv - data generated at UEF,
 Finland * pathDIA_Rd10_DR.csv - data generated at UEF, Finland *
 pathDDA_RPE65_KO.csv - data generated at UCI, US * pathDDA_P23H.csv - data
 generated at UCI, US * pathDDA_Rd10_DR.csv - data generated at UCI, US *
 pathDDA_Rd10_CLR.csv - data generated at UCI, US **Variables** * **ID**
 Identifier of the pathway or term (e.g., KEGG ID, GO term ID). *
 **Description** Name or description of the pathway or biological term
 (e.g., "Apoptosis", "Oxidative phosphorylation"). *
 **ProteinRatio** Proportion of differentially expressed proteins (DEPs)
 from your dataset found in the pathway: * **BgRatio** Background ratio —
 proportion of proteins associated with the pathway in the background
 (e.g., whole proteome): * **pvalue** Raw p-value from statistical test
 (e.g., hypergeometric test) indicating enrichment significance. *
 **p.adjust** Adjusted p-value (e.g., using Benjamini-Hochberg correction)
 to control for multiple comparisons. * **qvalue** False discovery rate
 (FDR) estimate — similar to p.adjust, used to assess significance while
 accounting for multiple testing. * **geneID of included DEPs** List of
 gene symbols or IDs for the differentially expressed proteins included in
 the pathway. * **Count** Number of DEPs from your dataset found in the
 pathway. This dataset describes characteristics of each group of samples,
 combined by genotype/group. **Files** * UCI_DDA_Rpe65-KO_cohort_1.csv -
 animals at UCI, USA * UEF_DIA_P23H_cohort_2.csv - animals at UEF, Finland
 * UCI_DDA_P23H_cohort_1.csv - animals at UCI, USA *
 UEF_DIA_rd10_DR_cohort_3.csv - animals at UEF, Finland *
 UCI_DDA_rd10_DR_cohort_2.csv - animals at UCI, USA *
 UCI_DDA_rd10_CLR_cohort_1.csv - animals at UCI, USA **Variables** *
 **Mouse ID** Unique identifier assigned to each individual mouse. * **File
 name** Name of the associated data file (e.g., raw data, image, or
 analysis file) for that mouse. * **Genotype** Genetic makeup of the mouse
 (e.g., WT for wild-type, or a specific mutation like P23H). * **Sex**
 Biological sex of the mouse (Male or Female). * **DOB** Date of Birth —
 the day the mouse was born. * **Euthanasia date** The date the mouse was
 euthanized (sacrificed for analysis). * **Euthanasia Method** The
 procedure used for euthanasia (e.g., CO₂ inhalation, cervical
 dislocation). * **Euthanasia age (days)** Age of the mouse at the time of
 euthanasia, calculated in days. * **Housing light condition** Description
 of the lighting environment in which the mouse was housed (e.g., 12:12
 light-dark cycle, constant darkness). ## Access information All files
 generated during the proteomic analysis | Dataset | Dataset identifier |
 Direct Link | | :---------------------------- |
 :---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | :--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | UCI\_DDA\_P23H\_cohort 1 | [PXD052547](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052547__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMhgrBoRU$) | [https://www.ebi.ac.uk/pride/archive/projects/PXD052547](https://urldefense.com/v3/__https:/www.ebi.ac.uk/pride/archive/projects/PXD052547__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrM0w8wbHE$) | | UCI\_DDA\_rd10\_DR\_cohort 2 | [PXD052549](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052549__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMeZQknLA$) | [https://www.ebi.ac.uk/pride/archive/projects/PXD052549](https://urldefense.com/v3/__https:/www.ebi.ac.uk/pride/archive/projects/PXD052549__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMpzrgaz0$) | | UCI\_DDA\_*Rpe65*-/- cohort 1 | [PXD052554](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052554__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMkCB7BOo$) | [https://www.ebi.ac.uk/pride/archive/projects/PXD052554](https://urldefense.com/v3/__https:/www.ebi.ac.uk/pride/archive/projects/PXD052554__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMehSOo5w$) | | UCI\_DDA\_rd10\_CLR\_cohort 1 | [PXD052555](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052555__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMGeUHWOM$) | [https://www.ebi.ac.uk/pride/archive/projects/PXD052555](https://urldefense.com/v3/__https:/www.ebi.ac.uk/pride/archive/projects/PXD052555__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMIoRHyEY$) | | UEF\_DIA\_rd10\_DR\_cohort 3 | [PXD052698](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052698__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMqSOnnTo$) | [https://panoramaweb.org/3IRD\_rd10.url](https://urldefense.com/v3/__https:/panoramaweb.org/3IRD_rd10.url__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMyeSYiDY$) | | UEF\_DIA\_P23H\_cohort 2 | [PXD052598](https://urldefense.com/v3/__http:/proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD052598__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMvpt1Vug$) | [https://panoramaweb.org/3IRD.url](https://urldefense.com/v3/__https:/panoramaweb.org/3IRD.url__;!!OLgoXmg!U4Q19XgPz9DYwXFrMNCASwqjtqDM3ueorMoq8Br8QWakCASR1FGLycqpR4IKI7eSszO9l9AjbBf0gLBn2WrMbADY64Q$) |