Specific subgroups of FruM neurons control sexually dimorphic patterns of aggression in Drosophila melanogaster

Abstract

A great challenge facing neuroscience is to understand how genes, molecules, cells, circuits, and systems interact to generate social behavior. Fruit flies (Drosophila melanogaster) offer a powerful model system to address questions of this magnitude. These animals display genetically specified, sexually dimorphic patterns of fighting behavior via sex-specific splicing of the fruitless gene. Here, we show that sexually dimorphic behavioral patterns displayed during aggression are controlled by specific subgroups of neurons expressing male forms of fruitless proteins (Fru(M)). Using the GAL4/UAS system to manipulate transformer expression, we feminized or masculinized different populations of neurons in fly nervous systems. With a panneuronal elav-GAL4 driver, male patterns of fighting behavior were transferred into females and female patterns into males. We screened 60 Gal4 lines that express the yeast transcription factor in different patterns in fly central nervous systems and found five that showed abnormal same-sex courtship behavior. The sexually dimorphic fighting patterns, however, were completely switched only in one and partially switched in a second of these lines. In the other three lines, female patterns of aggression were seen despite a switch in courtship preference. A tight correspondence was seen between Fru(M) expression and how flies fight in several subgroups of neurons usually expressing these proteins: Expression is absent when flies fight like females and present when flies fight like males, thereby beginning a separation between courtship and aggression among these neurons.

Fig. 1.

Behavioral patterns seen during courtship and aggression in wild-type and mutant flies. (A) Percentages of sexually dimorphic behavioral patterns seen during aggressive interactions in Canton-S, masculinized female and feminized male flies and in a Fru^F mutant background. Masculinized females (elav-GAL4/UAS-tra^IR) show male fighting patterns similar to Canton-S males, whereas feminized males (elav-GAL4/UAS-tra^F) show female fighting patterns similar to Canton-S females. No changes in fighting patterns were observed in masculinized females in a Fru^F background (elav-GAL4/UAS-tra^IR; fru^F/fru^F), confirming that Fru^M expression is responsible for the behavioral phenotypes we observed. “Lunge,” “Shove,” and “Headbutt” are the main components used to distinguish male-like and female-like patterns of fighting behavior. (B) Percentages of aggressive, courtship, and mixed interactions between pairs of flies. Both masculinized females and feminized males show increased same-sex courtship behavior (i.e., wing extension, singing, licking, or attempted copulation) when compared with wild-type flies. (C) Patterns of aggression shown by females masculinized by using five selected GAL4 drivers. With the l (3)31-GAL4 driver male fighting patterns are observed, others (c1003-, 60IIA-, 227-GAL4) show female fighting patterns, and the 1407-GAL4 driver shows intermediate patterns of fighting behavior. (D) Percentages of interactions masculinized females of the five selected GAL4 driver lines spend in aggression, courtship, and mixed interactions. (n) represents the total number of interactions.

Fig. 2.

An illustration of the Fru^M expression patterns in elav-GAL4/UAS-tra^IR masculinized female and elav-GAL4/UAS-tra^F feminized male pupal brains. Immunostaining of the posterior brain regions and adult brains are shown in SI Fig. 4. (A) Anterior brain regions of 48-h-old pupae of elav-GAL4/UAS-tra^IR masculinized females were dissected, fixed, and immunostained (see Methods) for Fru^M (green) and NC82 (magenta). (B). Schematic drawing of elav-GAL4/UAS-tra^IR masculinized female brains. (C) Anterior brain regions of 48-h-old pupae of elav-GAL4/UAS-tra^F feminized males. (D) Schematic drawing of elav-GAL4/UAS-tra^F feminized male brains. Reductions in Fru^M cell number are seen in fru-mAL and fru-aSP2 clusters in feminized males. (Scale bars, 50 μm.)

Fig. 3.

Colocalization of Fru^M (green) and β-galactosidase (magenta) staining reveals GAL4 expression at any time during development (see above text) in the l (3)31-GAL4 and 227-GAL4 lines. Images were obtained by crossing GAL4 drivers with w;UAS-FLP; Act5c(FRT.polyA)lacZ.nls reporter lines. White arrows indicate cell clusters that are Fru^M-positive but do not express GAL4. (Scale bars, 20 μm.)