|
Fig. 8
Posttranscriptional Hes6 protein oscillations.
(A) Wildtype (wt, left), her1 mutant (middle), and her7 mutant embryos at the 10-somite stage in situ stained for hes6 mRNA expression. (B) Intensity profile of hes6 mRNA staining in the embryos shown in (A), using the region boxed in black in the rightmost panel in (A). The origin of the graph corresponds to the end of the notochord. hes6 mRNA levels decay smoothly from posterior to anterior in wildtype, her1, and her7 mutant embryos. (C) Single confocal sections of 10-somite stage wildtype (left) and hes6 mutant embryos (right) immunostained with monoclonal antibodies raised against full-length Hes6 (yellow). Nuclei are counterstained with Hoechst 33342 (blue). Antibodies show immunoreactivity with a nuclear antigen in wildtype, but not in hes6 mutant embryos. Insets show magnified view of region boxed in red in the tailbud. (D) Widefield images of wildtype embryos at the 10-somite stage immunostained with Hes6 antibodies. Dynamic waves of Hes6 staining in the intermediate PSM (arrowheads) become apparent upon enhancing the contrast in this region (panels to the right). (E) Intensity profile of Hes6 protein staining for the embryos shown in (D) using the region boxed in white in the rightmost panel of (D). The origin of the graph corresponds to the end of the notochord. Waves of Hes6 protein levels manifest as local peaks in the staining profile (arrowheads). (F) Widefield images of her1 mutant (left) and her7 mutant (right) embryos at the 10-somite stage immunostained with Hes6 antibodies. Waves of Hes6 staining in the intermediate PSM can be observed upon contrast enhancement in her1 mutant embryos (arrowheads), but not in her7 mutant embryos. (G) Intensity profiles of Hes6 protein staining for the embryos shown in (F). Arrowheads point towards local peaks in the intensity trace indicative of protein oscillations, which can be detected for her1 mutant embryos (light and dark red traces), but not for the her7 mutant embryo (green trace). (A, C, D, F) Flat mount preparations, anterior to the top; scale bars, 50 μm.