Figure 1. xCPEB4 NTD is phosphorylated in response to progesterone.

(A) Schematic representation of xCPEB4 protein. The four protein fragments generated are: 1, from amino acid (aa) 1 to 202; 2, from aa 203 to 290; 3, from aa 291 to 409; and RBD, from aa 402 to 702. RRMs are shown in blue and the ZZ domain, in grey. (B) Lambda-phosphatase assay (λ) of oocytes overexpressing HA-xCPEB4 collected at the indicated times (VI, prophase I; GVBD, germinal vesicle breakdown; MII, metaphase II). Western blots with anti-HA antibody and anti-tubulin (as a loading control) are shown. C, control, corresponds to non-injected oocytes. (C) In vitro kinase assay of recombinant xCPEB4, full-length (FL) or the fragments 1, 2, 3 or RBD, using metaphase II oocyte extracts as the source of kinases (autoradiography, ³²P). (D) Time course of the in vitro kinase assay of recombinant xCPEB4, FL or fragments (1, 2 and 3), using oocyte extracts collected at the indicated times (autoradiography, ³²P). Histone H1 phosphorylation was used to follow Cdk1 activity. Three independent biological replicates were performed, with equivalent findings each time. See also Figure 1—figure supplement 1.

DOI: http://dx.doi.org/10.7554/eLife.19298.002

Figure 1—figure supplement 1. Loading controls for in vitro kinase assays.

(A) Loading control for Figure 1C. Upper panel corresponds to the Coomassie blue–stained gel (CBB with extract). In the lower panel, equivalent protein amounts used in the assay were loaded for a clearer loading control (CBB without extract). (B) Loading control for Figure 1D, in a Coomassie blue–stained gel.

DOI: http://dx.doi.org/10.7554/eLife.19298.003