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J M Nickerson, Felix L Struebing, Shanu Markand, Kevin Donaldson, Salma Ferdous, Jeffrey H Boatright, Eldon E Geisert; Alternative splicing in the retina in a large family of genes is regulated by a single trans locus. Invest. Ophthalmol. Vis. Sci. 2016;57(12):No Pagination Specified. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Initial observation identified a potentially novel form of gene regulation. The first exons in some genes are highly expressed in retinas of the C57BL/6J mouse, but expressed minimally in the retinas of the DBA/2J mouse, suggesting the use of alternative transcription start sites in both strains. This study tests the hypothesis that a distant single gene locus regulates this differential expression of these numerous alternative splicing events.
We examined retinas from 55 BXD lines by Affymetrix ST2.0 microarrays to find each exon expressed highly in one but not the other strain (C57BL/6J and DBA/2J). Highly variant exons with a Linkage Related Score (LRS) >50 (p <1.0 x 10-5) were used to identify trans quantitative trait loci (QTLs) that may modulate differential expression. Criteria used in candidate gene analysis within the trans-QTL included: retina expression, nuclear location, involvement with transcription or splicing mechanisms, and a sequence variation that would be causative.
177 exons were differentially expressed with an LRS > 50. Many of those were first exons, suggesting alternative transcription start sites. A predominant locus found on chromosome 4 modulated the differential expression of 98 of these 177 exons. This locus contains 21 intergenic sites and 41 genes with non-synonymous SNPs, any one of which might control alternative splicing in our group of genes. In this chromosome 4 locus, Znf593 appeared to be the best candidate. Znf593 mRNA was detected at high levels in the retina (16 fold above mean mRNA expression level), it had a strong cis-QTL (LRS = 77), and it contained a non-synonymous proline to arginine change at amino acid 47. Immunoreactivity for Znf593 protein was found abundantly in nuclei of the GCL and to a lesser extent in the INL and ONL in both C57BL/6J and DBA/2J mouse eyes.
A single amino acid change (proline to arginine) in Znf593 may control differential splicing for a large family of retinally-expressed genes. Znf593, a C2H2-type Zinc finger protein, interacts with Oct transcription factors, which together might modulate differential splicing or alternative transcription start sites observed between the C57BL/6J and DBA/2J retinas.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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