Unknown,Transcriptomics,Genomics,Proteomics

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Gene expression in retina and LGN of wild type and Chrnb2-/- mice

ABSTRACT: Mice lacking the beta 2 subunit (Chrnb2) of the neuronal nicotinic acetylcholine receptor display altered retinal waves and disorganized projections of the retinal ganglion cells to the lateral geniculate nucleus (LGN). mRNA populations from retinas and LGN from Chrnb2-/-and wild type (C57BL/6J) mice were compared at 4 days postnatal, when RGC segregation to the LGN begins in WT mice. Retinal mRNAs were also compared at adulthood. Using microarray hybridization, we identified transcripts which are differentially expressed between Chrnb2-/- and wild type animals in these two tissues at these two ages. mRNA was isolated from retina and LGN of three male littermates each of WT and Chrnb2-/- mice at P4. mRNA from retinas of two adult male littermates of each type was also examined.

ORGANISM(S): Mus musculus

SUBMITTER: Carol Rubin

PROVIDER: E-GEOD-22824 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:To investigate the retinal proteins associated with primary and secondary retinal ganglion cell (RGC) degeneration and explore their molecular pathways, SWATH label-free and target-based mass spectrometry was employed to identify the proteomes in various retinal locations in response to localized optic nerve injury. Unilateral partial optic nerve transection (pONT) was performed on adult Wistar rats and their retinas were harvested at 2 weeks later. To confirm the separation of primary and secondary RGC degeneration, immunohistochemistry of RNA binding protein with multiple splicing (RBPMS) and glial fibrillary acidic protein (GFAP) was performed on retinal whole-mounts. Retinal proteomes in the temporal and nasal quadrants were evaluated with high resolution hybrid quadrupole time-of-flight mass spectrometry (QTOF-MS), and SWATH-based acquisition, and their expression was compared to the corresponding retinal quadrant in contralateral control eyes and further validated by multiple reaction monitoring mass spectrometry (MRM-MS). A total of 3641 proteins (p<0.05, FDR<1%) were quantified by SWATH-MS. Bioinformatics data analysis showed that there were 35 up-regulated and 24 down-regulated proteins in the temporal quadrant while 19 and 5 proteins were up-regulated and down-regulated, respectively, in the nasal quadrant respectively (n=4, p<0.05; fold change ≥1.4 fold or ≤0.7). Six proteins were regulated in both the temporal and the nasal quadrants including CLU, GFAP, GNG5, IRF2BPL, L1CAM and CPLX1. Linear regression analysis indicated a strong association between the data obtained by SWATH-MS and MRM-MS (temporal, R2=0.97; nasal, R2=0.96). Gene ontology analysis reveals statistically significant changes in the biological processes and cellular components of primary RGC degeneration. The majority of the significant changes in structural, signaling, and cell death proteins were associated with loss of RGCs in the area of primary RGC degeneration. The combined use of SWATH-MS and MRM-MS methods detects and quantifies regional changes of retinal protein expressions after localized injury. Future investigation with this integrated approach will significantly increase understanding of diverse processes of progressive RGC degeneration from a proteomic prospective.

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Gene expression in retina and LGN of wild type and Chrnb2-/- mice

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