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Differences in corneal phenotypes between destrin mutants are due to allelic difference and modified by genetic background.

Kawakami-Schulz SV, Verdoni AM, Sattler SG, Ikeda A, Ikeda S - Mol. Vis. (2012)

Bottom Line: The goal of this study was to determine whether phenotypic differences are due to allelic differences between Dstn(corn1) and Dstn(corn1-2J), or are the result of genetic background effects.Actin accumulation, neovascularization, epithelial proliferation and inflammation in B6.Cg-Dstn(corn1) cornea are significantly reduced when compared to A.BY Dstn(corn1)cornea.Differences in the abnormal phenotypes of Dstn mutants result from allelic differences between Dstn(corn1) and Dstn(corn1-2J) .

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, University of Wisconsin, Madison, WI 53705, USA.

ABSTRACT

Purpose: Mutations in destrin (Dstn) cause corneal abnormalities in mice. A mutation, Dstn(corn1), results in corneal epithelial hyperproliferation, inflammation, and neovascularization in the A.BY background (A.BY Dstn(corn1)). Homozygosity for a point mutation, Dstn(corn1-2J), results in mild thickening of the corneal epithelium but no corneal neovascularization in a C57BL/6 (B6) background (B6 Dstn(corn1-2J)). The goal of this study was to determine whether phenotypic differences are due to allelic differences between Dstn(corn1) and Dstn(corn1-2J), or are the result of genetic background effects.

Methods: We generated two congenic (Cg) mouse lines, B6.Cg-Dstn(corn1) and A.BY.Cg-Dstn(corn1-2J), to compare to the original A.BY Dstn(corn1) and B6 Dstn(corn1-2J) lines. We performed immunohistochemistry to assay F-actin accumulation, neovascularization, proliferation, and inflammation. By western blot analysis we tested the expression of serum response factor (SRF), a known regulator of the Dstn(corn1) phenotype.

Results: The Dstn(corn1) mutation leads to neovascularization, hyperproliferation, and inflammation in the cornea of A.BY Dstn(corn1) as well as B6.Cg-Dstn(corn1) mice. We did not observe significant corneal neovascularization or hyperproliferation in either A.BY.Cg-Dstn(corn1-2J) or B6 Dstn(corn1-2J) mice. Actin accumulation, neovascularization, epithelial proliferation and inflammation in B6.Cg-Dstn(corn1) cornea are significantly reduced when compared to A.BY Dstn(corn1)cornea. SRF changes are consistent in Dstn(corn1) mutants, regardless of genetic background.

Conclusions: Differences in the abnormal phenotypes of Dstn mutants result from allelic differences between Dstn(corn1) and Dstn(corn1-2J) . Moreover, phenotypes of Dstn(corn1) mice are modified by genetic background, suggesting the existence of genetic modifiers. Protein analysis suggests that a genetic modifier affects phenotypic severity functionally downstream from or in a pathway independent from SRF. These data demonstrate that natural genetic variation affects phenotypic severity in Dstn(corn1) mice.

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Western blot analysis for SRF in Dstncorn1 mutant and WT cornea. A: SRF level is greater in Dstncorn1 mutant cornea compared to WT. B: When comparing the induction of SRF expression by the Dstncorn1 mutation (SRF increase in Dstncorn1 with respect to WT) in both the A.BY and B6 background, there is no significant difference in the increase of SRF between these genetic backgrounds. Error bars, SEM * denotes statistical significance resulting from t-tests. *p<0.05, **p<0.01, ***p<0.001, ns=nonsignificant.
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f6: Western blot analysis for SRF in Dstncorn1 mutant and WT cornea. A: SRF level is greater in Dstncorn1 mutant cornea compared to WT. B: When comparing the induction of SRF expression by the Dstncorn1 mutation (SRF increase in Dstncorn1 with respect to WT) in both the A.BY and B6 background, there is no significant difference in the increase of SRF between these genetic backgrounds. Error bars, SEM * denotes statistical significance resulting from t-tests. *p<0.05, **p<0.01, ***p<0.001, ns=nonsignificant.

Mentions: We had previously identified the transcription factor serum response factor (SRF) as a major contributor to Dstncorn1 phenotypes [2,6]. To assess the quantity of SRF in the Dstncorn1 mutants in both genetic backgrounds, we performed western blot analysis. We found that the level of SRF is higher in Dstncorn1 when compared WT in both backgrounds (Figure 6A). Additionally, statistical analysis reveals that the level of induction of SRF expression in Dstncorn1 mutants, that is, the increase relative to WT, is not significantly different between genetic backgrounds. (Figure 6B).


Differences in corneal phenotypes between destrin mutants are due to allelic difference and modified by genetic background.

Kawakami-Schulz SV, Verdoni AM, Sattler SG, Ikeda A, Ikeda S - Mol. Vis. (2012)

Western blot analysis for SRF in Dstncorn1 mutant and WT cornea. A: SRF level is greater in Dstncorn1 mutant cornea compared to WT. B: When comparing the induction of SRF expression by the Dstncorn1 mutation (SRF increase in Dstncorn1 with respect to WT) in both the A.BY and B6 background, there is no significant difference in the increase of SRF between these genetic backgrounds. Error bars, SEM * denotes statistical significance resulting from t-tests. *p<0.05, **p<0.01, ***p<0.001, ns=nonsignificant.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3298451&req=5

f6: Western blot analysis for SRF in Dstncorn1 mutant and WT cornea. A: SRF level is greater in Dstncorn1 mutant cornea compared to WT. B: When comparing the induction of SRF expression by the Dstncorn1 mutation (SRF increase in Dstncorn1 with respect to WT) in both the A.BY and B6 background, there is no significant difference in the increase of SRF between these genetic backgrounds. Error bars, SEM * denotes statistical significance resulting from t-tests. *p<0.05, **p<0.01, ***p<0.001, ns=nonsignificant.
Mentions: We had previously identified the transcription factor serum response factor (SRF) as a major contributor to Dstncorn1 phenotypes [2,6]. To assess the quantity of SRF in the Dstncorn1 mutants in both genetic backgrounds, we performed western blot analysis. We found that the level of SRF is higher in Dstncorn1 when compared WT in both backgrounds (Figure 6A). Additionally, statistical analysis reveals that the level of induction of SRF expression in Dstncorn1 mutants, that is, the increase relative to WT, is not significantly different between genetic backgrounds. (Figure 6B).

Bottom Line: The goal of this study was to determine whether phenotypic differences are due to allelic differences between Dstn(corn1) and Dstn(corn1-2J), or are the result of genetic background effects.Actin accumulation, neovascularization, epithelial proliferation and inflammation in B6.Cg-Dstn(corn1) cornea are significantly reduced when compared to A.BY Dstn(corn1)cornea.Differences in the abnormal phenotypes of Dstn mutants result from allelic differences between Dstn(corn1) and Dstn(corn1-2J) .

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, University of Wisconsin, Madison, WI 53705, USA.

ABSTRACT

Purpose: Mutations in destrin (Dstn) cause corneal abnormalities in mice. A mutation, Dstn(corn1), results in corneal epithelial hyperproliferation, inflammation, and neovascularization in the A.BY background (A.BY Dstn(corn1)). Homozygosity for a point mutation, Dstn(corn1-2J), results in mild thickening of the corneal epithelium but no corneal neovascularization in a C57BL/6 (B6) background (B6 Dstn(corn1-2J)). The goal of this study was to determine whether phenotypic differences are due to allelic differences between Dstn(corn1) and Dstn(corn1-2J), or are the result of genetic background effects.

Methods: We generated two congenic (Cg) mouse lines, B6.Cg-Dstn(corn1) and A.BY.Cg-Dstn(corn1-2J), to compare to the original A.BY Dstn(corn1) and B6 Dstn(corn1-2J) lines. We performed immunohistochemistry to assay F-actin accumulation, neovascularization, proliferation, and inflammation. By western blot analysis we tested the expression of serum response factor (SRF), a known regulator of the Dstn(corn1) phenotype.

Results: The Dstn(corn1) mutation leads to neovascularization, hyperproliferation, and inflammation in the cornea of A.BY Dstn(corn1) as well as B6.Cg-Dstn(corn1) mice. We did not observe significant corneal neovascularization or hyperproliferation in either A.BY.Cg-Dstn(corn1-2J) or B6 Dstn(corn1-2J) mice. Actin accumulation, neovascularization, epithelial proliferation and inflammation in B6.Cg-Dstn(corn1) cornea are significantly reduced when compared to A.BY Dstn(corn1)cornea. SRF changes are consistent in Dstn(corn1) mutants, regardless of genetic background.

Conclusions: Differences in the abnormal phenotypes of Dstn mutants result from allelic differences between Dstn(corn1) and Dstn(corn1-2J) . Moreover, phenotypes of Dstn(corn1) mice are modified by genetic background, suggesting the existence of genetic modifiers. Protein analysis suggests that a genetic modifier affects phenotypic severity functionally downstream from or in a pathway independent from SRF. These data demonstrate that natural genetic variation affects phenotypic severity in Dstn(corn1) mice.

Show MeSH
Related in: MedlinePlus