Transcript Supplementary Figure 4 - PowerPoint (327 KB )

Supplemental 4: C. elegans Rbx-1 is unlikely a component of the RPM-1/FSN-1 protein complex
a
b
rbx-1 RNAi treatment
(against ZK287.5)
-
+
N2
N2
IP antibody
juIs58
56 kDa
N2
juIs58;
juIs58 fsn-1
N2
juIs58;
juIs58 fsn-1
FSN-1
Nonspecific
30 kDa
30 kDa
20kDa
20 kDa
RBX-1
RBX-1
b-tubulin
56 kDa
b-tubulin
10% lysate input
56 kDa
Panel a: A polyclonal antibody raised against yeast RBX1 (provided by M. Tyers, University of Toronto) was tested against
the C. elegans total protein lysates. Adult wild-type animals (N2) were fed with bacteria expressing dsRNA against
ZK287.5 (the C. elegans homologue of RBX1) for three days, and the protein lysates were probed with RBX1 antibodies
at 1:2000 dilution. Compared with lysates from untreated animals, a band corresponding to the predicted molecular
weight of C. elegans RBX-1 protein was consistently decreased/abolished in three sets of experiments. This band most
likely corresponds to the C. elegans RBX-1 protein.
Panel b: Immunoprecipition from wild-type (N2), RPM-1::GFP (juIs58) and fsn-1;RPM-1::GFP(juIs58) lysates by antibodies
against GFP. FSN-1 specifically associates with RPM-1::GFP, while RBX-1 does not. The same condition as in
Figure 2 was used for the co-immunoprecipitation experiment.