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Fig 6. Phosphorylation of Nemp1. A. Developmental analysis for modified Xl_Nemp1. Xenopus embryos were injected with mRNA for Xl_Nemp1-HA and collected at the indicated stages (St.). Lysates were subjected to western blotting with anti-HA or β tubulin antibody (loading control). uc, uninjected control. B. In vitro alkaline phosphatase assay of Xl_Nemp1. Lysates were prepared at the late blastula stage (stage 9). Xl_Nemp1-HA was immunoprecipitated by anti-HA antibody and treated with (+) or without (-) calf intestinal alkaline phosphatase (CIAP). C. In vitro alkaline phosphatase assay of Mm_Nemp1. Lysates were prepared at the mid blastula stage (stages 8–8.5), and treated with (+) or without (-) λ protein phosphatase (λPP). D. Co-IP of Mm_Ran with phosphorylation site mutants of Mm_Nemp1. mRNA for Myc-tagged Mm_Ran was injected into Xenopus embryos with mRNA for HA-tagged Mm_Nemp1, its alanine mutant (5SA), or its glutamic acid mutant (5SE). Injected embryos were collected at the mid blastula stage and lysed with lysis buffer B. This data is the same as lanes 1, 2, 7, and 8 shown in S3 Fig Black arrowheads, modified forms. Image published in: Shibano T et al. (2015) Image reproduced on Xenbase with permission of the publisher and the copyright holder. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View

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