Segregation distortion induced by wild-type RanGAP in Drosophila

Abstract

Segregation Distorter (SD) is a meiotic drive system in Drosophila that causes preferential transmission of the SD chromosome from SD/SD(+) males owing to the induced dysfunction of SD(+) spermatids. The key distorter locus, Sd, is a dominant neomorphic allele encoding a truncated, but enzymatically active, RanGAP (RanGTPase-activating protein) whose nuclear mislocalization underlies distortion by disrupting the Ran signaling pathway. Here, we show that even wild-type RanGAP can cause segregation distortion when it is overexpressed in the male germ line or when the gene dosage of a particular modifier locus is increased. Both manipulations result in substantial nuclear accumulation of RanGAP. Distortion can be suppressed by overexpression of Ran or Ran guanine nucleotide exchange factor (RanGEF) in the male germ line, indicating that the primary consequence of nuclear mislocalization of RanGAP is reduction of intranuclear RanGTP levels. These results prove that segregation distortion does not depend on any unique properties of the mutant RanGAP encoded by Sd and provide a unifying explanation for the occurrence of distortion in a variety of experimental situations.

Figure 1

Immunolocalization of RanGAP in primary spermatocytes. Confocal images show staining with anti-RanGAP antibody (A, C, and D) or anti-HA (B) antibody on the Left and merged images with propidium iodide staining of chromatin on the Right. (A) Wild-type males. (B) Males expressing Sd-RanGAP tagged with an HA (influenza hemagglutinin) epitope (see ref. 18). (C) Males expressing β2-RanGAP. (D) Males carrying two doses of E(SD). Scale bar = 5 μm.

Figure 2

Western blot analysis of RanGAP expression in testes. Each lane contains the total protein from two pairs of testes boiled in SDS sample buffer, separated electrophoretically, transferred onto poly(vinylidene difluoride)-plus membrane, and probed with anti-RanGAP antibody. (A) Expression of RanGAP from the β2-tubulin promoter results in an approximate 20-fold increase in RanGAP expression. Lanes 1 and 2, Two independent transgenic lines expressing β2-RanGAP; lanes 3 and 4, two independent transgenic lines expressing β2-RanGAP R87A; lane 5, Cantons-S (control). (B) Expression of RanGAP is not increased in males carrying extra doses of E(SD). Lane 1, Males carrying two doses of E(SD); lane 2, males carrying one dose of E(SD); lane 3, Canton-S (control) males lacking E(SD).