Control of male courtship behavior by the fruitless gene.

Our previous studies led us to propose that a sex determination regulatory gene, fruitless (fru), is responsible for building the potential for male sexual behavior into the CNS during development in D. melanogaster. Our lab, in collaboration with the laboratories of Jeff Hall (Brandeis) and Barbara Taylor (Oregon State University) are continuing our studies of courtship behavior with the long term aims of understanding: (1) how the potential for male courtship behavior is established in the central nervous system (CNS) during development, and (2) how the cells subserving male courtship behavior function together to insure the ordered manifestation of the events comprising this behavior.
We have shown that wild-type fru function is required for all, or nearly all, aspects of male sexual behavior, from the initial recognition of a potential mate, through the transfer of bodily fluids and the duration of copulation. The sex determination hierarchy, in which fru functions, specifies all somatic sexual differences between males and females. The genes acting immediately above fru in the sex determination hierarchy direct the sex-specific splicing of the transcripts of the distal (P1) fru promoter to produce female- and male-specific fru mRNAs (Figure 1). Genetic analysis indicates that it is the P1-derived male-specific FRU proteins (FRUM) whose functions are needed for male sexual behavior. FRUM proteins are three BTB domain Zn-finger transcription factors that have different Zn-finger pairs due to the use of alternative 3' exons.
Genomic map of the fruitless gene.  Top:  Coordinates of fru exons derived from cDNA sequences mapped onto the genomic sequence.  Black rectangles represent exons and inverted triangles represent the positions of p-element insertion mutations.  Middle:  Diagram of cDNA classes with introns denoted by lines connecting the exons (black rectangles).  Alternative splicing at the sex-specific sites is shown by the open rectangle, which is used only in females.  Alternative internal 32 nt, 15 nt and 148 nt micro-exons are shown without connections to other exons.  Question marks indicate where the complete array of exons used in a particular transcript class is unknown.  Bottom:  Proteins encoded by the fru P1-derived mRNAs.
That fru acts during development to establish the potential for male sexual behavior is suggested by the patterns of expression of male-specific P1 products. The FRUM proteins are expressed almost exclusively in the CNS, where they are first detected in a small number of cells at the end of the larval period. Expression is maximal about two days into the pupal period when ca. 1700 cells (2% of the CNS) express FRUM proteins; this time coincides with the period of major morphogenetic events that shape the adult CNS (Figure 1). Cells expressing the FRUM proteins are not localized to one part of the CNS. They are scattered throughout the brain and VNC, most frequently found in small groups of cells, and less frequently as single cells. There are approximately 20 groups of FRUM neurons (Fig. 2). Some of these groups are in specific regions of the CNS previously shown to be involved in particular steps of courtship behavior. In addition, a number of regions containing FRUM neurons had not been previously implicated in courtship. Classification of the types of neurons in which fru is expressed-based on the locations, size and morphologies of their cell bodies-provides insight into fru's function. Most cells expressing these products are in higher order sensory or motor neuropils and have morphologies suggesting that they are either local circuit neurons or interganglionic interneurons. Few of the cells expressing the male-specific fru products appear to be either sensory or motor neurons. One exception is a group of serotonergic FRUM cells in the abdominal ganglion that provide the sole innervation of much of the internal male genitalia. These findings suggest that male sexual behavior is the result of sensory information entering the CNS via largely sex-nonspecific sensory systems, and being processed by sex-specific (fru-specified) circuitry in higher order neuropils. This information is then used to execute the particular steps of courtship behavior by modulating largely sex-nonspecific motor systems.
Figure 2.  Anti FRUM staining of wild-type male CNS.
The findings that FRUM proteins are expressed in only a small portion of the CNS and are necessary for all, or nearly all, aspects of male sexual behavior were provocative and pleasing. They suggested fru provided a handle for understanding the developmental, genetic, molecular, and neuronal bases of male courtship behavior. Yet there is a formidable gap between the two aspects of fru about which we have substantial information: (1) the properties and expression patterns of its products and (2) fru's phenotypic effects on male sexual behavior. It is on bridging this gap that our current research is focused. We believe the key to gaining an understanding of how the potential for a complex behavior is built into the CNS will be to go from the gene (fru) to the properties of the cells in which it functions and from those cells to behavior. Thus it is on the developmental origins of these neurons, the characteristics that distinguish them, and the roles they play in adult male sexual behavior that we are focused. We are addressing the following topics.
  1. What are the behavioral roles of individual groups of FRUM neurons?
  2. To begin to understand the functions of the various groups of FRUM neurons we are (A) identifying what types of neurons they are, (B) asking how homologously positioned groups of P1 fru-expressing neurons in males and females differ, and (C) for selected groups of cells, determining their neuronal connections and synaptic partners to begin to elucidate the neuronal circuitry underlying male courtship behavior.
  3. How is the pattern of the FRUM neurons generated during development? We are determining when the FRUM cells are born during development, whether FRUM cells found in clusters are clonally related to one another, and the reasons why some groups of cells expressing the P1 promoter are found in only one sex.
  4. How do the FRUM proteins function? The FRU proteins are putative BTB Zn-finger transcription factors. We are (A) determining whether FRU proteins function as homo- or heterodimers with other BTB domain proteins, investigating the role of the unique 101 amino acid N-terminus of the male-specific FRU proteins, and (B) identifying genes that are regulated by FRU proteins.
  5. Finally, we are asking: Is fru is both necessary and sufficient for male courtship behavior?

 

Publications on fruitless and male courtship behavior from the Baker, Hall and Taylor laboratories collaboration.

Original research papers:

Belote, J. M. and B. S. Baker (1987). On the establishment and maintenance of an inate action pattern by genes regulating sex determination in Drosophila. Proc. Natl. Acad. Sci. USA, 84: 8026-8030.

Taylor, B. J., Villella, A., Ryner, L. C., Baker, B. S., and Hall, J. C. (1994). Behavioral and Neurobiological implications of sex-determining factors in Drosophila. Devel. Genetics 15: 275-296.

Ryner, L. C., Goodwin, S. F., Castrillon, D. H., Anand, A., Villella, A., Baker, B., Hall, J. C., Taylor, B. J., and Wasserman, S. A. (1996). Control of male sexual behavior and sexual orientation in Drosophila by the fruitless gene. Cell, 87:1079-1089.

Villella, A., Gailey, D. A., Berwald, B., Ohshima, S., Barnes, P. T., and Hall, J. C. (1997). Extended reproductive roles of the fruitless gene in Drosophila melanogaster revealed by behavioral analysis of new fru mutants. Genetics 147: 1107-1130.

Heinrichs, V., Ryner, L.C. and Baker, B.S. (1998). Regulation of sex-specific selection of fruitless 5' splice sites by transformer and transformer-2. Mol. Cell. Biol. 18: 450-458.

Goodwin S.F., Taylor, B.J., Villella, A., Foss, M., Ryner, L. C., Baker, B. S., and Hall, J. C., (2000) Aberrant splicing and altered spatial expression patterns in fruitless mutants of Drosophila melanogaster. Genetics 154: 725-45.

Lee, G., Foss, M., Goodwin, S., Carlo, T., Taylor, B. J., and Hall, J. C. (2000). Spatial, temporal and sexually dimorphic expression patterns of the fruitless gene in the Drosophila CNS. J. Neurobiol. 43: 404-426.

Lee, G., and Hall, J. C., (2000). A newly uncovered phenotype associated with the fruitless gene of Drosophila melanogaster: aggression-like head interactions between mutant males. Behav. Genet. 30: 263-275.

Lee, G., Villella, A., Taylor, B. J., Hall, J. C. (2001). New reproductive anomalies in fruitless-mutant Drosophila males: Extreme lengthening of mating durations and infertility associated with defective serotonergic innervation of reproductive organs. J. Neurobiol. 47: 121-149.

Lee, G. and Hall, J.C. (2001). Abnormalities of male-specific FRU protein and serotonin expression in the CNS of fruitless mutants in Drosophila. J. Neurosci. 21: 513-526.

Anand, A., Villella, A., Ryner, L. C., Carlo, T., Goodwin, S. F., Song, H.-J., Gailey, D. A., Hall, J. C., Baker, B. S., and Taylor, B. J., (2001). The sex determination gene fruitless , in addition to controlling male sexual behaviors, has sex-nonspecific vital functions. Genetics, 158: 1569-1595.

Song, H.-J.,. Goodwin , S. F., Reynaud, E., Carlo, T., Billeter, J.-C., Eric P. Spana, E. P., Perrimon, N., Baker, B. S., and Taylor, B. J., (2002). The fruitless gene is required for the proper formation of axonal tracts in the embryonic CNS of Drosophila. Genetics, 162: 1703-1724.

Song, H.-J. and Taylor, B. J. (2003). The fruitless gene is required to maintain neuronal identity in evenskipped-expressing neurons in the embryonic CNS of Drosophila. J. Neurobiol., in press.

Reviews:

Baker, B. S., Taylor, B. T., and Hall, J. C., (2001) Are complex behaviors specified by dedicated regulatory genes? Reasoning from Drosophila. Cell 105: 13-24.