Fig. 6

Catalase Overexpression in RPCs Is Sufficient to Trigger Their Premature Differentiation

(A) Early catalase overexpression in rx2-expressing retinal progenitor cells (RPCs) leads to a decrease of H₂O₂ levels in RPCs at 24 h postfertilization (hpf) as revealed by the HyPer transgene reporting the endogenous levels of H₂O₂ of UAS:catalase-E2A-tagRFP (UAS:cat-E2A-tagRFP) injected in the double transgenic Tg(ubi:HyPer);Tg(rx2:gal4) embryos (white arrow and red dash line).

(B) Normalized quantification of H₂O₂ levels in catalase-expressing (RFP-positive) clones at 24 hpf (catalase, n = 21 RPCs) and neighboring cells RFP-negative (control, n = 13 RPCs). p value < 0.0001, depicted as ^∗∗∗∗, unpaired Student’s t test.

(C) Early catalase overexpression by injection of UAS:catalase-E2A-tagRFP in Tg(rx2:gal4) RPCs led to the premature differentiation of their deriving cells as revealed by the late photoreceptor marker Zpr1 (white arrowhead, Zpr1+ Ph) in contrast to control UAS:tagRFP-injected RPCs.

(D) Quantification of Zpr1-positive cells in UAS:tagRFP control clones and UAS:cat-E2A-tagRFP expressing clones revealed a significant increase in the number of Zpr1-positive cells in clones that expressed catalase (n = 7 retinae for UAS:tagRFP control and n = 13 retinae for UAS:cat-E2A-tagRFP, p value = 0.0255, Mann-Whitney U test). Medians for both conditions are shown (0 for UAS:tagRFP control and 2 for UAS:cat-E2A-tagRFP).

(E) UAS:catalase-E2A-tagRFP construct was injected in double transgenic Tg(rx2:gal4);Tg(atoh7:GFP) embryos at 1-cell stage and the effect on atoh7:GFP expression was assessed at 26 hpf, before the normal onset of atoh7 expression. An induction of atoh7:GFPexpression in the RFP-positive clones expressing catalase in the dorsal retina could be observed, whereas no GFP could be detected in UAS:tagRFP-control-injected retinae (n = 10 retinae). Scale bars, 10 μm in (A) and (C) and 20 μm in (E).