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Fig. 3. Example of a nonobvious disease model revealed by phenologs: a yeast model of angiogenesis. (A) The sets of 8 genes (considering only mouse/yeast orthologs) associated with mouse angiogenesis defects and 67 genes associated with yeast hypersensitivity to the hypercholesterolemia drug lovastatin significantly overlap, suggesting that the yeast gene set may predict angiogenesis genes. This prediction was verified in Xenopus embryos for eight genes (three from literature support and five based upon vascular expression patterns) (Fig. S3) and studied in detail for the case of the transcription factor sox13. (B) sox13 is expressed in developing Xenopus vasculature, as measured by in situ hybridization (also Fig. S4). (C) Morpholino (MO) knockdown of sox13 induces defects in vasculature, measured using in situ hybridization versus the vasculature markers erg (defects observed in 31 of 49 animals tested) or agtrl1 (12 of 19 animals tested) (Fig S5). Such defects are rare in untreated control animals and five base pair mismatch morpholino (MM) knockdowns (0 of 22 control animals tested with agtrl1, 2 of 46 tested with erg; 5 of 28 MM animals tested with erg). (D) Hemorrhaging (white arrows) is apparent in stage 45 Xenopus embryos because of dysfunctional vasculature following sox13 morpholino knockdown (12 of 50 animals tested; two also showed unusually small hearts with defective morphology; Right: magnification of yellow boxed region in Middle), but is rare in control animals (1 of 45 tested untreated animals, 1 of 22 sox13-MM knockdown animals tested). All phenotypes in Figs. 3 and 4 are significantly different from controls by χ2 tests (P < 0.001). (E) In an in vitro human umbilical vein endothelial cell model of angiogenesis, knockdown of human SOX13 by siRNA disrupts tube formation (an in vitro model for capillary formation) to an extent comparable to knockdown of a known effector of angiogenesis (HOXA9) and significantly more than untreated cells or cells transfected with an off-target (scrambled) negative control siRNA. (Scale bar, 100 μm.) |
