GAL4 enhancer trap targeting of the Drosophila sex determination gene fruitless.

The fru4 allele of the sex determination gene fruitless is induced by insertion of a P[lacZ,ry+] enhancer trap element. This insert also acts to disrupt expression of the fru P1 promoter derived male-specific proteins, consequently impairing male courtship behavior. fru4 maps less than 2 kb upstream of the fru P3 promoter, whose function is essential for viability. We replaced this insert with a GAL4 element, P[GAL4,w+], recovering two lines with insertions in opposite orientations at the locus, one of which demonstrated fru-specific mutant phenotypes. Reporter expression of these lines recapitulated that of P3- and P4-derived proteins which, when correlated with a developmental and tissue specific survey of fru promoters' activities, uncovered a previously unsuspected complexity of fru regulation. These novel fru alleles provide the tools for manipulation of fru-expressing cells, allowing the consequent effects to be related back to specific fru functions and the regulatory units controlling these activities.

Dornan AJ ，Gailey DA ，Goodwin SF 《GENESIS》

Characterization of Drosophila fruitless-gal4 transgenes reveals expression in male-specific fruitless neurons and innervation of male reproductive structures.

The fruitless (fru) gene acts in the central nervous system (CNS) of Drosophila melanogaster to establish male sexual behavior. Genetic dissection of the locus has shown that one of the fru gene's promoter, P1, controls the spatial and temporal expression of male-specific FruM proteins critical to determining stereotypical male sexual behavior. By using the Gal4-expression system, we show that a 16-kb fragment of the fru P1 promoter's 5' regulatory region drives the expression of Gal4 in a subset of FruM-expressing neurons within both the pupal and adult CNS. Colocalization of FruM and a Gal4-responsive reporter shows that the fru(P1)-gal4 fusion construct generates expression in both previously characterized FruM-expressing neurons as well as within cells of both the CNS and the peripheral nervous system that have not been demonstrated as FruM-expressing. Gal4-expressing neurons are shown to innervate abdominal organs directly relevant to fru function; specifically, the muscle of Lawrence (MOL) and the male internal reproductive organs. Innervations of the latter are shown to originate from identified FruM-serotonergic neurons. Furthermore, we show that the MOL neuromuscular junction is sexually dimorphic. Finally, we describe Gal4 expression in neurites innervating male reproductive structures that are hypothesized to be targets of fru function. Isolation of the regulatory sequences controlling the expression of fru in the CNS, therefore, provides a potent tool for the manipulation of FruM-expressing neurons and for understanding the cellular basis of Drosophila reproductive behavior.

Billeter JC ，Goodwin SF 《JOURNAL OF COMPARATIVE NEUROLOGY》

A gene necessary for normal male courtship, yellow, acts downstream of fruitless in the Drosophila melanogaster larval brain.

The fruitless (fru) gene is a member of the Drosophila melanogaster somatic sex determination genetic pathway. Although it has been hypothesized that the primary function of fru is to regulate a genetic hierarchy specifying development of adult male courtship behavior, genes acting downstream of fru have not yet been identified. Here we demonstrate that the yellow (y) gene is genetically downstream of fru in the 3(rd)-instar larval brain. Yellow protein is present at elevated levels in neuroblasts, which also show expression of male-specific FRU proteins, compared to control neuroblasts without FRU. A location for y downstream of fru in a genetic pathway was experimentally demonstrated by analysis of fru mutants lacking transcription of zinc-finger DNA binding domains, and of animals with temporal, spatial, or sexual mis-expression of male-specific FRU. A subset of fru and y mutants is known to reduce levels of a specific behavioral component of the male courtship ritual, wing extension, and FRU and Yellow were detected in the general region of the brain whose maleness is necessary for development of that behavior. We therefore hypothesized that ectopic expression of Yellow in the 3(rd)-instar brain, in a y null background, would rescue low levels of wing extension and male competitive mating success, and this was found to be the case. Overall, these data suggest that y is a downstream member of the fru branch of the D. melanogaster sex determination hierarchy, where it plays a currently unknown role in the development of adult male wing extension during courtship.

Drapeau MD ，Radovic A ，Wittkopp PJ ，Long AD ... -  《journal of neurobiology》

fruitless gene is required to maintain neuronal identity in evenskipped-expressing neurons in the embryonic CNS of Drosophila.

The fruitless (fru) gene acts sex-nonspecifically in the development of the embryonic central nervous system (CNS) and has sex as well as sex-nonspecific functions in the development of the adult CNS. In the embryo, sex-nonspecific fru mRNAs and proteins are widely expressed during neurogenesis and present in both neurons and glia. To assess whether the fru gene played any role in fate determination of neuronal precursors and neurons, we examined the development of Eve-positive (Eve(+)) GMCs and neurons in fru mutants. In fru mutant embryos in which most or all fru transcripts were eliminated, the normal complement of Eve(+) neurons was present initially, but some neurons were unable to maintain their Eve-expression. Concomitantly, a subset of Eve(+) neurons also showed inappropriate expression of the glial marker, reversed polarity. In addition, neurons that normally do not express Eve became Eve(+) in these fru mutants. These defects were rescued in fru mutant embryos expressing specific fru transgenes under the control of the sca-GAL4 and elav-GAL4 drivers. These phenotypic analyses and rescue experiments provide evidence that one of the sex-nonspecific functions of the fru gene is the maintenance of neuronal identity rather than establishment of a neuron's initial fate.

Song HJ ，Taylor BJ 《journal of neurobiology》

fruitless splicing specifies male courtship behavior in Drosophila.

Demir E ，Dickson BJ 《Cell》