Fig. 4.

Wisp colocalizes and interacts with Aub. a, b Immunostaining of UASp-GFP-Aub nos-Gal4 stage 10 oocytes, also stained with DAPI (blue) (a) and 0–2 h-embryos (b), with anti-GFP and anti-Wisp. Posterior poles are shown in the bottom panels in a, and in b. Quantification of colocalization in the germ plasm using the Manders coefficient is shown in b. Scale bars: 30 μm in a top panels, and 10 μm in a bottom panels and in b. c Immunostaining of UASp-GFP-Aub/nos-Gal4 0–2 h-embryos with anti-GFP and either anti-CCR4 (top panels), or anti-Smg (bottom panels). Scale bars: 10 μm. d Co-immunoprecipitation of Wisp with GFP-Aub in UASp-GFP-Aub/nos-Gal4 0–2 h-embryos. w ¹¹¹⁸ 0–2 h-embryos were used as negative control (Mock). Immunoprecipitation was with anti-GFP (GFP IP) either in the presence (+) or the absence (−) of RNase A. e Co-immunoprecipitation of Aub with Wisp in wild-type, csul and tud mutant 0–2 h-embryos. Immunoprecipitation was with anti-Wisp in the presence of RNase A. Bound proteins were detected using western blots with anti-Wisp and anti-Aub; inputs correspond to protein extracts before IP in d, e. f GST pull-down assays between GST-Wisp and HA-Aub. Constructs and interactions are shown in the table. HA-tagged Aub fragments were revealed using western blot with anti-HA. Inputs correspond to 1:10 of in vitro-synthetized HA-Aub fragments before pull-down. GST alone was used as a negative control. GST and GST-recombinant proteins used in each pull-down are shown (bottom panels). Arrowheads indicate full-length recombinant proteins