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insxenopus kidney 

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Experiment details for ins

Romaker D et al. (2014) Assay

MicroRNAs are critical regulators of tuberous sclerosis complex and mTORC1 activity in the size control of the Xenopus kidney.

Gene Clone Species Stages Anatomy
ins.L laevis NF stage 33 and 34 pronephric kidney

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  Fig. S5. Expression analysis of Insulin/Insulin-like growth factor (Igf) signaling. Whole-mount in situ hybridizations and paraplast sections thereof comparing igf1 (A–B′′), igf3 (C–D′′), insulin receptor (insr) (E–F′′), insulin receptor substrate 1 (G–H′′), igf2 (I), insulin (ins) (K), igf receptor (igfr) (M), and insulin-related receptor (insrr) (N) at stages 34 (A, C, E, G, I, and K) and 38 (B–B′′, D–D′′, F–F′′, H–H′′, M, and N). (J and L) In situ hybridizations on paraplast sections of stage 42 Xenopus embryos for igf2 and insulin. Insets show sense control.

Gene Clone Species Stages Anatomy
ins.L laevis NF stage 37 and 38 pronephric kidney

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  Fig. 3. Insulin/Igf2 activate mTORC1 in the proximal tubules. (A–B′′) Whole-mount in situ hybridizations and paraplast sections thereof showing expression of Insulin or Igf2 in the proximal tubules of stage 38 Xenopus embryos. (C–D′) Phospho-Insulin/IGF receptor (pInsR/IgfR) immunofluorescence analysis of uninjected controls and embryos injected with Ins/Igf2-MOs at stage 40. C and D show pInsR/IgfR staining only; C′ and D′ show merged images with DAPI (blue) and ECL (green) to visualize nuclei and proximal tubules, respectively. Note the apical activation of the Insulin/IGF receptor present in control embryos, which is lost in tubules lacking Insulin and Igf2. (E–H′) pAkt and pS6 immunofluorescence analysis of uninjected controls and Ins/Igf2-MO–injected embryos at stage 42. E, F, G, and H show pAkt or pS6 staining only; E′, F′, G′, H′ show merged images with DAPI (blue) and 3G8 (green) to visualize nuclei and proximal tubules, respectively. Tubular structures in C, D, E, F, G, and H are outlined by yellow dotted lines. (I–K) 3G8 whole-mount immunohistochemistry and total proximal tubular cell counts of control embryos and Insulin/Igf2 morphants at stage 42. Number of embryos analyzed and the SD is indicated in the individual bars. *P < 0.005. (L) Stage 42 cell counts from uninjected (blue) and Ins/Igf2-MO–injected embryos (red) superimposed on the scatterplot of Fig. 1F (gray) demonstrate that impaired growth is caused by decreased numbers of mitotic cells. The fact that the pronephroi of the Insulin/Igf2 morphants shift to the right indicates that proliferation is not completely abolished but reduced to a baseline level.