Fig. 1. GLD-2 genes and proteins in Drosophila. (A) Schematic of GLD-2 proteins from different species. Accession numbers are NP_572766 for Wisp, NP_651012 for CG5732-encoded protein, NP_491842 for C. elegans, AAT98005 for Xenopus and NP_776158 for human. The regions showing homology are in gray and black; percentage similarity with Wisp in the central (including catalytic) domain and in the PAP/25A domain are indicated. The mutation D1031A in the catalytic domain that precludes poly(A) polymerase activity, and the point mutation in wisp^12-3147, are shown. (B) Schematic of wisp (CG15737) locus and mutant. Black boxes are exons. The arrow indicates the transcription start site. The three cDNAs LD18468, RE03648 and RE14825 were sequenced. RE03648 and RE14825 are full-length and start and end at identical nucleotides, whereas LD18468 is incomplete at both ends. The P-element (not drawn to scale) in wisp^KG5287 is shown. The insertion site was verified by sequencing. (C) Western blots with anti-Wisp, showing Wisp expression in females and ovaries and the lack of protein in wisp^KG5287, wisp⁴⁰ and wisp⁸⁹ mutant ovaries. Protein extracts were from 0.4 (lane 1) or 0.2 (lanes 2, 9) wild-type ovaries, from one (lanes 3-6) or 0.2 (lanes 7, 8) mutant ovaries, from 0.3 male (lane 10) or 0.1 female (lane 11), and from 20 wild-type embryos (right panel). -tubulin was used as a loading control. (D) Immunostaining of ovaries with anti-Wisp, showing cytoplasmic expression (green) throughout oogenesis and the lack of expression in wisp^KG5287 mutant ovaries. Nuclei are visualized with DAPI (blue). Anterior is oriented towards the left.