Abstract
Purpose :
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.
Methods :
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.
Results :
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.
Conclusions :
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.