Fig. 4. PAPC and Nlk1 function cooperatively during gastrulation regulation. A, Quantification of gastrulation defects in stage 35 Xenopus embryos injected at four-cell stage dorsal-equatorially with the indicated MOs (6 ng/embryo). Total amount of MO injected in each embryo was adjusted to 12 ng/embryo with control (Cont.) MO. B, Quantification and representative images of AC explants from embryos injected animally at the four-cell stage with indicated MOs (5 ng/embryo, total MO amounts adjusted with Cont. MO). ACs were excised at stage 8–9, elongation was induced with 50 ng/ml Activin, and samples were scored after 14 h. C, ATF2 luciferase reporter assay in stage 12 embryos injected at the eight-cell stage animal-dorsally with indicated MOs (5 ng/embryo, total MO amounts adjusted with Cont. MO). The reporter was activated by injection of wnt11 mRNA (250 pg/embryo). The graph shows the mean±SD of three independent experiments (* p<0.05 compared with Wnt11 induced control). RLU: relative light units. D, Immunoblot analysis of endogenous PAPC in gastrula stage Xenopus embryos. Embryos were injected at the four-cell stage with the indicated MOs (10 ng/embryo) and mRNA (300 pg/embryo). β-Tubulin was used as loading control. E, Quantification of AC explants from embryos injected animally at the four-cell stage with indicated MOs (12 ng/embryo) and mRNA (300 pg/embryo). ACs were excised at stage 8–9 and elongation was induced with 50 ng/ml Activin, and samples were scored after 14 h. The legend of the graph is exactly as in B. F, Brachet's cleft formation of sagittally sectioned stage 11 embryos injected with the indicated MOs (12 ng/embryo) and synthetic mRNA (300 pg/embryo). Categorization of the cleft for quantification in the graph on the right is shown at the bottom of the representative images. The blastopore (posterior) is marked by black arrowheads, red and green arrowheads indicate the anterior and posterior end of the Brachet´s cleft, respectively.
Image published in: Kumar R et al. (2017)
Copyright © 2017. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License.
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