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Dev Biol
2007 Oct 15;3102:341-7. doi: 10.1016/j.ydbio.2007.08.005.
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Cyclin E2 is required for embryogenesis in Xenopus laevis.
Gotoh T
,
Shigemoto N
,
Kishimoto T
.
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In mammalian cells, E-type cyclins (E1 and E2) are generally believed to be required for entry into S phase. However, in mice, cyclin E is largely dispensable for normal embryogenesis. Moreover, Drosophila cyclin E plays a critical role in cell fate determination in neural lineages independently of proliferation. Thus, the functions of cyclin E, particularly during early development, remain elusive. Here, we investigated the requirement for E-type cyclins during Xenopus embryogenesis. Although cyclin E1 has been reported as a maternal cyclin, inhibition of its translation in the embryo caused no serious defects. We isolated a Xenopus homologue of human cyclin E2, which was zygotically expressed. Sufficient inhibition of its expression led to death at late gastrula, while partial inhibition allowed survival. These observations indicate distinct roles for Xenopus cyclins E1 and E2, and an absolute requirement of cyclin E2 for Xenopus embryogenesis.
Fig. 4.
Xcyclin E1 is dispensable for development after MBT. (A) A morpholino oligo for Xcyclin E1 (XcycE1-MO) was targeted to the underlined sequence. The box indicates the start codon. (B) One-cell stage embryos were uninjected or injected with 80 ng of either control MO (Cont-MO) or XcycE1-MO, cultured, collected at indicated times, and analyzed by Western blot with anti-Xcyclin E1 or anti-phospho-Cdc2-Tyr15 (Cdc2-P) antibodies. (C) The Xcyclin E1 signals shown in panel B were quantified by phosphoimager. (D) Embryos as in panel B were photographed when uninjected control embryos reached the indicated stages. Note that neural tube was not still closed in XcycE1-MO-injected embryos when uninjected embryos reached stage19. Scale bar, 250 μm.
Fig. 5.
Sufficient loss of Xcyclin E2 is lethal during late gastrulation. (A) Two types of morpholino oligos for Xcyclin E2 (XcycE2-MO1 and -MO2) were targeted to underlined sequences, respectively. The box indicates the start codon. (B) One-cell stage embryos were uninjected or injected with 80 ng of either Cont-MO, XcycE2-MO1 or XcycE2-MO2, cultured, collected when uninjected embryos reached at stages 7 and 11, and analyzed by Western blot with anti-Xcyclin E2 (top) or control anti-MAPK antibodies (bottom). Asterisk indicates non-specific bands. (C) Embryos as in panel B were photographed when uninjected embryos reached the indicated stages. Arrows indicate blastopore. Scale bar, 250 μm. (D) Embryos as shown in panel B were inspected for embryonic death when uninjected embryos reached stage 15. The error bar represents the standard deviation (SD) from triplicate samples.
Fig. 6.
Sufficient loss of both Xcyclin E1 and E2 is embryonic lethal at late gastrulation. (A) One-cell stage embryos were uninjected or injected with 160 ng of Cont-MO, 80 ng of XcycE1-MO or XcycE2-MO1, or each 80 ng of both XcycE1-MO and XcycE2-MO1, cultured, collected when uninjected embryos reached stages 7 (early blastula) and 11.5 (late gastrula), and analyzed by Western blot with anti-Xcyclin E1 (top), anti-Xcyclin E2 (middle) or control anti-MAPK antibodies (bottom). Asterisk indicates non-specific bands. (B) Embryos as in panel A were photographed when uninjected embryos reached the indicated stages. Arrows indicate blastopore. Scale bar, 250 μm. (C) Embryos as in panel A were inspected for embryonic death when uninjected embryos reached stage 15. The error bar represents the SD from triplicate samples.
