Depletion of <i>cxcr4a</i> impairs G1/S transition and Foxj1a protein expression in DFCs.

Abstract

(A–B) Representative confocal sections of BrdU-positive DFCs and KV cells at the indicated stages (A). Dorsal views with anterior on the top. Scale bar, 20 μm. The percentage of BrdU-positive cells among GFP-positive DFCs and KV cells were quantified from the indicated embryo numbers in three independent experiments, and the group values are expressed as the mean ± SD (B). Underlying data can be found in S1 Data. (C-D) BrdU incorporation experiments showed reduced proliferating DFCs in cxcr4aum20 mutants at the 75% epiboly stage. Scale bar, 20 μm. Statistical data from three independent experiments are shown in (D). Student t test, ***P S1 Data. (E–I) Depletion of cxcr4a inhibits the G1/S transition in DFCs. Representative confocal sections of WT and cxcr4a-deficient Tg(sox17:GFP;EF1α:mKO2-zCdt1(1/190)) embryos at the bud and 3-somite stages are shown in (E) and (G). Dorsal views with anterior to the top. Scale bar, 20 μm. The percentage of mKO2-positive KV cells were quantified from three independent experiments (F and H). The average numbers of KV cells in WT embryos and cxcr4aum20 mutants at the 3-somite stage are shown in (I). The significance of differences compared with the WT group was analyzed with the Student t test, **p p S1 Data. (J–K) cxcr4a deficiency down-regulates zFoxj1a protein expression levels. WT and cxcr4a-deficient Tg(sox17:GFP) embryos were harvested at the 75% epiboly and bud stages and then subjected to western blot analysis (J) and immunostaining (K) with the indicated antibodies. Scale bar, 20 μm. BrdU, bromodeoxyuridine; DFC, dorsal forerunner cell; Ef1α, eukaryotic translation elongation factor 1α; ep, epiboly; Foxj1a, forkhead box j1a; GFP, green fluorescent protein; KV, Kupffer’s vesicle; mKO2, monomeric Kusabira Orange2; sox, SRY-box transcription factor; Tg, transgene; WT, wild-type; zCdt1, zebrafish chromatin licensing and DNA replication factor 1; zFoxj1a, zebrafish Foxj1a; α-Tubulin, acetylated tubulin.</p