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Fig. 6. XPACE4 depletion affects the processing of a specific subset of TGFß proteins. (A) Maturation of Xnr2 is reduced in XPACE4-depleted embryos. Western blots from control (Un) and XPACE4-depleted [P(-)] whole embryos and blastocoel fluids overexpressing Xnr2-HA mRNA (200 pg) show the mature form of Xnr2-HA (26 kDa) is reduced in whole embryo homogenates of XPACE4-depleted embryos. In the blastocoel fluid of XPACE4-depleted embryos, a significant reduction in mature Xnr2 level and accumulation of the unprocessed (57 kDa) and an intermediate form (33 kDa) are detected. (B) Maturation defect of Xnr2 in XPACE4-depleted embryos is rescued by injection of XPACE4 mRNA (150 pg). In whole embryos, the level of mature Xnr2 is rescued by XPACE4 mRNA injection. In the blastocoel fluid the unprocessed and intermediate forms are rescued by XPACE4 mRNA injection. (C) Maturation of Xnr1 is reduced in XPACE4-depleted embryos expressing 200 pg of Xnr1-HA mRNA. The mature form of Xnr1-HA (24 kDa) is reduced in whole embryo homogenates of XPACE4-depleted embryos. In the blastocoel fluid, a significant reduction in mature Xnr1 level and accumulation of the unprocessed form (57 kDa) are detected. (D) Maturation of Xnr3 is reduced in XPACE4-depleted embryos expressing 300 pg of Xnr3-HA mRNA. The mature form of Xnr3-HA (22 kDa) is reduced in whole embryo homogenates of XPACE4-depleted embryos. In the blastocoel fluid, a significant reduction in mature Xnr3 level is detected. (E) Maturation of Xnr5 is not affected in XPACE4-depleted embryo lysates or blastocoel fluids. Xnr5-HA mRNA (100 pg) is injected. (F) Maturation of ActivinB is not affected in XPACE4-depleted embryos. A very low level of mature ActivinB-HA (15 kDa) is more concentrated in the blastocoel fluid both in controls and XPACE4-depleted whole embryos. However, there is no significant change in levels of mature (15 kDa) or unprocessed forms (54 kDa) of ActivinB-HA. ActivinB-HA (400 pg) mRNA is injected. (G) Maturation of Derrière is not affected in XPACE4-depleted embryos. There is no significant change in levels of mature (21 kDa) or unprocessed (50 kDa) forms of Derrière-HA, either in whole embryo homogenates or in the blastocoel fluid of XPACE4-depleted embryos. Derrière-HA mRNA (400 pg) is injected. (H) Maturation of Vg1 is reduced in XPACE4-depleted embryos. The unprocessed Vg1 as a doublet (46 and 44 kDa), and intermediate form (35 kDa) are easily detected in whole embryo lysates and the blastocoel fluid. The mature form (18 kDa) is reduced in the blastocoel fluid of XPACE4-depleted embryos. Vg1-HA mRNA (600 pg) is injected. ({alpha}-tubulin is used as a loading control.)





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