Phenotype Image Detail

Image

Caption

Altered expression of genes involved in vascular development in Igs1^{tm11(CAG-Bgeo,-Edn2)Nat}/Igs1⁺ Tg(Six3-cre)69Frty/0 (Z/Edn2;Six3-Cre) retinas. (A) Sections of central and peripheral P6 control and mutant retinas stained with Toluidine Blue. In the mutant retina, patent blood vessels are observed on the vitreal face of the central retina (arrowheads), but the peripheral retina is avascular. Except for thinning in the periphery, the morphology of the mutant retina is largely normal. (Scale bar: 100 um.) (B) Microarray hybridization with total retina RNA from four biologically independent pairs of P8 control and mutant samples. In the volcano plot, probe sets showing more than twofold reduction in hybridization intensity in the mutant sample and a P value < 0.05 are shown in green, and probe sets showing more than twofold increase in hybridization intensity in the mutant sample and a P value < 0.05 are shown in red. Blue symbols represent Apln and Vegfa probe sets, which show an approximately twofold increase in hybridization intensity and P values < 0.05. In situ hybridization for these two transcripts is shown in D and G. Because of the presence of nonspecific background signal, microarray hybridization intensity changes typically underestimate the actual fold change in RNA abundance. (C) Expression profiling using RNAseq. The data presented is from a single sequencing experiment with RNA from P5 control and mutant retinas. The ratio (on a log2 scale) of read counts for individual transcripts from control vs. mutant libraries is shown for all genes with more than twofold ratios (i.e., log2 > 1) and P values < 0.05. Transcripts highlighted in B are highlighted here. (D-I) In situ hybridization to retina sections from P6 control and mutant retinas. Probes for tip cell markers are (D) Apln, (E) Dll4, and (F) Ang2. In D, Apln transcripts are present only on the vitreal face of the retina. They are localized to a narrow annulus of tip cells in the periphery of the control retina (bracket in boxed region enlarged in Inset in D, Left) but broadly distributed in surface ECs near the center of the mutant retina (D, Right). E-I show sections of central retina. In E, Dll4 transcripts are not detectable in the control retina but localize to ECs on the vitreal surface of the mutant retina. In F, Ang2 transcripts are present in a subset of cells in the INL (probably horizontal cells) in both control and mutant retinas; Ang2 transcripts are absent from ECs in the control retina and present in ECs on the vitreal surface of the mutant retina. In G, Vegfa transcripts are present in the retinal pigment epithelium (RPE) and INL in both control and mutant retinas (with a stronger INL signal in the latter); Vegfa transcripts are not detected on the vitreal surface of the control retina but are abundant in astrocytes in mutant retinas. In H and I, GS-lectin staining after in situ hybridization to mutant retinas shows (H) Apln transcripts colocalizing with GS-lectin in ECs and (I) Vegfa transcripts in astrocytes interdigitating with ECs. ISH, in situ hybridization. (Scale bars: D, 500 um; E-G, 100 um; H and I, 50 um.) (J-O) In situ hybridization to retina flat mounts from P6 control and mutant retinas. Green, GS-lectin; red, in situ hybridization signal. Boxed regions are enlarged as Insets. The in situ hybridization signal in H-O consists of purple alkaline phosphatase reaction product; it has been false colored red for illustrative purposes. (Scale bar: 500 um.) PlxnD1, Plexin-D1.

Copyright

This image is from Rattner A, Proc Natl Acad Sci U S A 2013 Oct 1;110(40):E3830-E3839. Copyright 2013 National Academy of Sciences, U.S.A. J:201133

Associated Alleles

Symbol	Name
Igs1^tm11(CAG-Bgeo,-Edn2)Nat	intergenic site 1; targeted mutation 11, Jeremy Nathans
Tg(Six3-cre)69Frty	transgene insertion 69, Yasuhide Furuta

Associated Genotypes

Allelic Composition	Genetic Background
Igs1^tm11(CAG-Bgeo,-Edn2)Nat/Igs1^+ Tg(Six3-cre)69Frty/0	involves: 129S1/Sv * 129X1/SvJ * C57BL/6 * DBA/2