FIG. 2.
Molecular genetics of gus and Fsn. (A) Schematic representation of the gus locus and its transcripts (not to scale). Insertion sites of the PBac elements used in this study are indicated. (B) RT-PCR amplifications specific for the RA class and gus-RB transcripts indicate that both mRNAs are expressed in wild-type ovaries (lane A). gusf07073 impairs expression of all tested transcripts (lane B), while guse00456 in trans to gusf07073 (lane C) or homozygous (lane D) affects only gus-RB. Positions of the primer pairs used are indicated in panel A; exact sequences are available upon request. A fragment of cul-1 mRNA was amplified as a control. (C) An antibody raised against recombinant Gus recognizes a ∼30-kDa protein that is reduced in gusf07073 ovaries, corresponding most likely to the 281-amino-acid-long isoform of Gus (GusS). The blot was also probed with anti-α-tubulin antibodies (α-Tub) as a loading control. (D) Transgenic GusS::HA efficiently coimmunoprecipitates Vas. A peptide containing an intact DINNN motif moderately interferes with this reaction, while a corresponding peptide bearing the N188A substitution within the DINNN motif does not have this effect. (E) Eyes from flies bearing the gusf07073, guse00456, and gusf04367 insertions. The mini white marker that tags the PBac shows position effect variegation in its expression. (F) In trans to the deficiency Df(2R)nap1, the gusf07073, guse00456, and gusf04367 mutations cause defects in the wing, including blisters, folded wings and ectopic vein material. (G) Hoyer's embedded wings derived from Df(2R)nap1/gusf07073 flies. (H) Schematic representation of the Fsn locus (not to scale). The insertion site of PBac Fsnf06595 is indicated. (I) RT-PCR amplifications specific for the Fsn-RA and Fsn-RB transcripts indicating expression of both isoforms in wild-type ovaries (lane A). In Df(2R)Exel7124/Fsnf06595 ovaries (lane B) these mRNAs are not expressed at detectable levels. (J) Eggs produced by gusf07073/gusf07073 mothers. Many of these eggs collapse.