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. Author manuscript; available in PMC: 2017 Mar 7.
Published in final edited form as: Dev Cell. 2016 Mar 7;36(5):562–571. doi: 10.1016/j.devcel.2016.02.010

Fig. 1. Identification of cytoplasmic isoforms encoded by Drosophila Rbfox1.

Fig. 1

(A) Schematic of Drosophila germline cyst differentiation. Germline stem cells (GSC) give rise to cystoblasts (CBs) that then divide four times to form 16-cell cysts. These divisions are incomplete and cells remain connected through cytoplasmic bridges called ring canals (magenta). During these divisions the fusome (green) branches (region 1 and 2A). Fusomes are specialized organelles that become increasingly more branched up until the 16-cell cyst stage, when they begin to undergo degradation (de Cuevas and Spradling, 1998). The fusome is labeled with the adducin-like protein Hu li tai shao (Hts). After germ cells complete their mitotic divisions, the fusome degrades (region 2B). Persistence of fusomes typically indicates a defect in germ cell differentiation. Cytoplasmic Sex lethal (Sxl) is expressed in GSCs and stays on until the 2-cell cyst stage. Rbfox1 is first observed in 4-cell cysts, with a peak of expression at 8-cell and Region 2A 16-cell cyst stage. Rbfox1 levels subsequently decrease. (B) Schematic of the Rbfox1 locus showing that Rbfox1-RF and Rbfox1-RN both use an alternative splice site that results in exclusion of a nuclear localization signal (NLS) contained in the other annotated isoforms. (C) A wild-type germarium stained for Rbfox1 (magenta), the fusome marker Hts (green) and DNA (blue). Regions 1, 2A, 2B and 3 are indicated. (C′) Rbfox1 staining alone. The bracket marks germ cells that express high levels of cytoplasmic Rbfox1. Small arrows mark somatic follicle cells that express nuclear Rbfox1. (D) vasa-gal4>UAS-HA::Rbfox1-RF (vasa-gal4>HA::Rbfox1-RF) germarium stained for the HA-tagged transgene (magenta) and DNA (blue). (E–F) Germaria from (E) vasa-gal4/CyO; Rbfox1e03440/Df (vasa-gal4>Rbfox1e03440/Df), (F) vasa-gal4>UAS-HA::Rbfox1-RF; Rbfox1e03440/Df (vasa-gal4>HA::Rbfox1-RF; Rbfox1e03440/Df) and (G) nanos-gal4; UAS-Rbfox1-RFRNAi (nanos-gal4>Rbfox1-RFRNAi) females stained for Hts (green) and DNA (blue). (H) Rbfox1dsRed.1 homozygous mutant germarium stained for Rbfox1 (magenta), Hts (green) and DNA (blue). The bracket marks cysts that lack high levels of cytoplasmic Rbfox1. Arrows mark Rbfox1dsRed.1 mutant somatic cells that retain nuclear Rbfox1. Scale bars represent 10 μm.