XB-IMG-85312
Xenbase Image ID: 85312
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FIG. 7. GREUL1 functions as an E3 ubiquitin ligase and the RING
domain is necessary for both E3 activity and XAG-1 induction. (A)
Comparison of the GST fusion constructs to wild type GREUL1.
Abbreviations: TM, transmembrane domain; RING, RING finger
domain; wt, wild type; *, point mutation (cysteine replaced by
glycine); the triangle represents the signal peptide cleavage site. (B)
Upper panel: Western-blot with FLAG antibody visualizing FLAGtagged
ubiquitin chains, ranging from 25 to 250 kDa. The Apc2/Apc11 (Apc2/11) complex was used as a control. Lower panel:
Western blot with GST antibody showing the actual amount of
GSTC-term and GST C1C2 added to the above reactions. 10, 30,
and 100 ng of total purified GSTC-term and 10, 30, 100, 300, and
900 ng of total purified GST C1C2 was used in each reaction. Only
30 and 100 ng of GSTC-term were able to catalyze ubiquitination.
The size of the protein band is 49 kDa. (C, D) A functional RING
domain is necessary for XAG-1 induction. 1 ng of either wild type
GREUL1 (D) or C1C2 GREUL1 (C) was injected into two-cell
embryos and stained by in situ hybridization for XAG-1. Ectopic
XAG-1 dots are not apparent in the C1C2 GREUL1-injected
embryos. Image published in: Borchers AG et al. (2002) Copyright © 2002. Image reproduced with permission of the Publisher, Elsevier B. V.
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