Fig. 6. Expression of an uncleavable form of Ovhts disrupts the transition from fusome to RC. (A) Schematic of the location of the four 20-amino acid deletions. (B) Western blot of deletion constructs 1 through 4 expressed in S2 cells, probed with htsRC antibody. Ovhts-3 protein is not cleaved. (C) Ovary extracts from flies expressing either Ovhts::GFP or Ovhts-3::GFP blotted with htsRC antibody. Whereas only a cleavage product is present in Ovhts::GFP extract (open arrow), Ovhts-3::GFP is only present in a full-length band (arrowhead). (D-D") Germaria from flies expressing Ovhts::GFP show a GFP signal on RCs but not on fusome (arrow D'), and 1B1 localization only to fusome and not to RCs (arrow D"). (E-E") Flies expressing Ovhts-3::GFP show localization of GFP (E') and 1B1 (E") to both the fusome and RCs (arrows). (F,G) Stage 2 egg chambers expressing either Ovhts::GFP or Ovhts-3::GFP stained with 1B1 antibody (red). (F) RCs show localization of Ovhts::GFP in tight clear rings that do not have 1B1 staining. (G) RCs show a colocalization of 1B1 staining and Ovhts-3::GFP fluorescence on RCs (yellow). Additionally, RCs are malformed. (G',G") Magnification of boxed region in G shows that rings are not round, appear thicker and less organized, and can be fully occluded with fusome material that does (arrow) or does not (arrowhead) include Ovhts-3::GFP. Scale bars: C,D, 20 µm; F,G, 10 µm; G',G", 5 µm.