QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

Help viewing high resolution images Return to article

Fig. 2. Vls interacts physically with Csul. (A) Vls interacts with Csul in vivo. Ovarian protein extracts of homozygous vls³ females producing both TAP-Csul and HA-Vls (lanes 1 and 2), or only HA-Vls (lane 3), were directly loaded on a gel (one tenth of total input, lane1) or immunoprecipitated with Sepharose IgG (lanes 2 and 3). Following separation by SDS-PAGE electrophoresis, bound-HA-Vls was detected by immunoblotting using anti-HA antibodies. (B) Reciprocally, Vls was immunoprecipitated using anti-HA antibodies and bound Csul was detected by immunoblotting using alkaline phosphatase-conjugated anti-rabbit antibodies. (C) Binding of the C terminus of Vls to Csul. Full-length GST-Vls or derivatives were purified from bacterial extracts and incubated with S•Tag-Csul. Bound S•Tag-Csul (75 kDa) was separated by SDS-PAGE electrophoresis and detected by immunoblotting using alkaline phosphatase-conjugated S proteins. Input: one-tenth of protein extract was loaded on the gel. The amount of used GST-fusion proteins was evaluated by SDS-PAGE followed by Coomassie staining (lower panel). Representation of the GST-Vls constructs used for the mapping and summary of results are indicated on the right. The WD-repeats and the putative Csul-binding domain of Vls are indicated by grey and green boxes, respectively. (D) The C-terminal region of Csul interacts with Vls. Full-size S•Tag-Csul or derivatives were synthesized in vitro and incubated with full-size GST-Vls. Following separation by SDS-PAGE electrophoresis the bound S•Tag-Csul proteins were detected by immunoblotting using alkaline phosphatase-conjugated S proteins. Left panel: input S•Tag-Csul proteins. The arrow indicates an endogenous protein synthesized in the reticulocyte system and reactive to alkaline phosphatase-conjugated S proteins. White squares indicate the position of the different S•Tag-Csul proteins. Middle panel: S•Tag-Csul proteins bound to GST-Vls are indicated by white squares. Representation of S•Tag-Csul constructs used for the mapping and summary of the results are indicated in the right panel. The putative Vls-binding domain of Csul is depicted in blue.

Return to article