Maternal and Zygotic Sex-Specific Gene Interactions in DROSOPHILA MELANOGASTER

Sex-lethal (Sxl) is a vital, X-chromosome gene involved in Drosophila sex determination. The most striking aspect of the phenotype of daughterless (da), an autosomal maternal-effect mutation, may be explained by effects on the functioning of the Sxl gene in the zygote. In this paper, new aspects of interactions between various combinations of Sxl and da alleles are explored in order to understand better the complex da phenotype. The study focuses on the relationship between maternal and zygotic da+ gene functions, and on the relationship between aspects of the da phenotype that are sex-specific and aspects that are not. The SxlM#1 allele, which counteracts the female-specific maternal effect of da, is shown to have no effect on two other aspects of the da phenotype (one maternal, one primarily zygotic) that are not sex-specific. The female-lethal da maternal effect is shown to kill daughters even when the progeny are entirely wild-type with respect to da. Recessive mutant alleles of the two genes can interact synergistically when both are heterozygous with their wild-type alleles, disrupting the development of most of the daughters. Surprisingly, even a deficiency of the da+ locus can produce a dominant, temperature-sensitive, female-lethal maternal effect. A new class of subliminal Sxlf alleles is described. These spontaneous mutations can confuse analysis of both da and Sxl if their presence is not appreciated. Finally, conditions are described that facilitate the study of the Enhancer of daughterless mutation.     

The Drosophila sex determination gene daughterless has different functions in the germ line versus the soma

As a regulator of the female-specific gene Sxl, da+ provides an essential maternal component in the control of sex determination and dosage compensation; nevertheless, neither the maternal nor zygotic phenotypes of the original mutant da allele is sex-specific. Here we clarify the role of da+ in Drosophila development, finding: this sex determination gene is indeed pleiotropic; zygotic functioning of da+ is essential in both sexes for somatic cell development, but not for germ cell development; da female sterility results from a somatic, rather than germ-line, defect; and expression of da+ in the maternal germ line is required only for daughters in the subsequent generation, as expected for a specific regulator of Sxl+. These conclusions follow from the characterization of new da null alleles isolated by a selection for defects in maternally acting positive regulators of Sxl.     

The interaction between daughterless and sex-lethal in triploids: a lethal sex-transforming maternal effect linking sex determination and dosage compensation in Drosophila melanogaster

Regulation of Drosophila sex determination and X-chromosome dosage compensation in response to the X-chromosome/autosome (X/A) balance of the zygote is shown to require proper functioning of both the da+ gene in the mother and the Sxl+ gene in the zygote. Previous studies led to the hypothesis that zygotic Sxl+ alleles are differentially active in females (XXAA) vs males (XYAA) in response to the X/A balance, and that maternal da+ gene product acts as a positive regulator in this connection. Sxl+ activity was proposed to impose the female developmental sequence on cells which would follow the male sequence in its absence. Important predictions of this proposal are verified. This study focuses primarily on the phenotype of triploid intersexes (XXAAA, X/A = 0.67). They are shown here to survive effects of da and Sxl mutations that would be lethal to diploids. The ambiguous X/A signal of intersexes normally causes them to develop as phenotypic mosaics of male and female tissue. Loss of maternal da+ or zygotic Sxl+ gene function shifts their somatic sexual phenotype to the male alternative. A gain-of-function mutation at Sxl has the opposite effect, imposing female development regardless of the maternal genotype with respect to da. It also reduces their rate of X-linked gene expression. The effects of a duplication of Sxl+ resemble those of the constitutive Sxl allele, but are less extreme. The role of these genes in the process of X-chromosome dosage compensation is inferred indirectly from the strict dependence of the mutations' lethal effects on the X/A balance in haploids, diploids, and triploids, and more directly from the effects of the mutations on the phenotypes of the X-linked neomorphic mutations, Bar and Hairy-wing. The relationship of da+ and Sxl+ gene functions to those of other sex-specific lethal loci in D. melanogaster, and to sex determination mechanisms in other species, is discussed.     

Functional Changes Associated with Structural Alterations Induced by Mobilization of a P Element Inserted in the Sex-lethal Gene of Drosophila

Genetic analysis of rearrangements within the multifunctional sex determining gene Sex-lethal has allowed correlation of changes in specific functions with DNA alterations. Rearrangements were isolated by mobilization of a P element which is on the 5' side of the gene, at coordinate 0. Previous work has shown that rearrangements associated with alterations in Sxl gene function are found within an 11-kb region between coordinates-11 and 0. Here it is shown that insertion of foreign DNA, per se, at coordinate 0 is compatible with wild-type gene function. However, deletion of sequences on either side of this point generates a mutant phenotype. Deletions extending distally beyond coordinate -6.5 kb result in a null phenotype, whereas smaller distal deletions or proximal deletions eliminate only some Sxl functions.     

Sex determination in Drosophila: the X-chromosomal gene liz is required for Sxl activity.

In Drosophila, females require products of the gene Sxl for sex determination, dosage compensation and fertility. I show here that the X-chromosomal gene liz, located in 4F1 to 4F11 and previously called fs(1)1621, provides maternal and zygotic functions necessary for Sxl activity in germ line and soma. In XX animals, the mutation SxlM1 which was reported to express the female-specific functions of Sxl constitutively can rescue all phenotypes resulting from lack of liz product. XY animals carrying SxlM1 and lacking maternal or zygotic liz activity survive as males with some female traits. A stock was constructed in which the females are liz SxlM1/liz SxlM1 and males liz SxlM1/Y. This shows that SxlM1 is not truly expressed constitutively in animals with an X:A ratio of 0.5, but requires activity of liz for initiation or maintenance.     

Evidence That Sisterless-a and Sisterless-B Are Two of Several Discrete ``numerator Elements'''' of the X/a Sex Determination Signal in Drosophila That Switch Sxl between Two Alternative Stable Expression States

The primary signal for Drosophila sex determination is the number of X chromosomes relative to the number of sets of autosomes. The present report shows that the numerator of this X/A signal appears to be determined by the cumulative dose of a relatively limited number of discrete X-linked genetic elements, two of which are sisterless-a and sisterless-b. This discovery regarding the nature of the sex determination signal grew out of previous studies of both the likely X/A signal target (the feminizing switch gene, Sex-lethal) and two positive regulators of that target gene (sis-a and daughterless). Combinations of genetic perturbations in these three genes had been shown to have synergistic effects. A model proposed in part to account for these interactions generated a large variety of strong predictions for sex-specific synergistic interactions that would be diagnostic for X/A numerator elements and could distinguish them from other components of the sex determination system. All these predictions, as well as other predictions for X/A numerator elements, are shown here to be fulfilled. The most compelling observations involve sexually reciprocal viability effects of duplications of wild-type genes: combinations of sis-a+, sis-b+ and/or Sxl+ duplications are lethal to males but rescue females from the otherwise lethal effects of changes in other components of the sex determination machinery. The many interactions described here illustrate an important principle that may seem counter-intuitive: perturbations of the sex determination signal for Drosophila generally will not appear to affect adult sexual phenotype. This principle follows from the fact that Sxl is involved in dosage compensation as well as sex determination, and from important aspects of the nature and timing of Sxl's regulation both by the X/A signal and by Sxl's own products (positive autoregulation). These factors mask potential effects on adult sexual differentiation by causing the premature death of cells and/or individuals. The fact that the vast array of results presented here conform to this principle is strong evidence in favor of a "binary state" model for Sxl regulation by the X/A signal. This model is favored over an alternative "multiple state" hypothesis that was proposed by others in a different study of the X/A signal. In that same study it was concluded that region 3E8-4F11 of the X chromosome contained especially potent X/A numerator elements.(ABSTRACT TRUNCATED AT 400 WORDS)     

Parental effects and phenotypic characterization of mutations that affect early development in Caenorhabditis elegans

Genetic tests for parental effects were performed on 24 temperature-sensitive embryonic-lethal mutants of the nematode Caenorhabditis elegans. For 21 of these mutants, maternal expression of the wild-type allele is sufficient for embryonic survival, regardless of the embryo's genotype. For 11 of these 21 mutants, maternal expression of the wild-type allele is necessary for embryonic survival (strict maternals). For the remaining 10, either maternal or embryonic expression is sufficient for survival (partial maternals). One mutant shows a paternal effect; that is, a wild-type extragenic sperm function appears to rescue homozygous mutant embryos. Similar parental-effect tests were performed on 11 larval-lethal mutants. In 4 mutants, 1 of which blocks as late as the second larval stage after hatching, maternal contributions still can rescue mutant larvae. The remaining 3 embryonic lethals and 8 larval lethals show no parental effects; that is, zygotic expression of the wild-type allele is necessary and sufficient for embryonic survival. Temperatureshift experiments on embryonic-lethal embryos showed that all but 1 of the strict maternal mutants are temperature sensitive only before gastrulation. One of the partial maternal mutants is temperature sensitive prior to gastrulation, suggesting that some zygotic genes can function early in embryogenesis. At the nonpermissive temperature, 7 of the strict maternal mutants either show cleavage abnormalities in early divisions or stop cleavage at less than 100 cells, or both.     

The master switch gene sex-lethal promotes female development by negatively regulating the N-signaling pathway

Notch (N) signaling is used for cell-fate determination in many different developmental contexts. Here, we show that the master control gene for sex determination in Drosophila melanogaster, Sex-lethal (Sxl), negatively regulates the N-signaling pathway in females. In genetic assays, reducing Sxl activity suppresses the phenotypic effects of N mutations, while increasing Sxl activity enhances the effects. Sxl appears to negatively regulate the pathway by reducing N protein accumulation, and higher levels of N are found in Sxl(-) clones than in adjacent wild-type cells. The inhibition of N expression does not depend on the known downstream targets of Sxl; however, we find that Sxl protein can bind to N mRNAs. Finally, our results indicate that downregulation of the N pathway by Sxl contributes to sex-specific differences in morphology and suggest that it may also play an important role in follicle cell specification during oogenesis.     

Genetic analysis of slipper/mixed lineage kinase reveals requirements in multiple Jun-N-terminal kinase-dependent morphogenetic events during Drosophila development

Mixed lineage kinases (MLKs) function as Jun-N-terminal kinase (JNK) kinase kinases to transduce extracellular signals during development and homeostasis in adults. slipper (slpr), which encodes the Drosophila homolog of mammalian MLKs, has previously been implicated in activation of the JNK pathway during embryonic dorsal epidermal closure. To further define the specific functions of SLPR, we analyzed the phenotypic consequences of slpr loss and gain of function throughout development, using a semiviable maternal-effect allele and wild-type or dominant-negative transgenes. From these analyses we confirm that failure of dorsal closure is the null phenotype in slpr germline clones. In addition, there is a functional maternal contribution, which can suffice for embryogenesis in the zygotic null mutant, but rarely suffices for pupal metamorphosis, revealing later functions for slpr as the maternal contribution is depleted. Zygotic null mutants that eclose as adults display an array of morphological defects, many of which are shared by hep mutant animals, deficient in the JNK kinase (JNKK/MKK7) substrate for SLPR, suggesting that the defects observed in slpr mutants primarily reflect loss of hep-dependent JNK activation. Consistent with this, the maternal slpr contribution is sensitive to the dosage of positive and negative JNK pathway regulators, which attenuate or potentiate SLPR-dependent signaling in development. Although SLPR and TAK1, another JNKKK family member, are differentially used in dorsal closure and TNF/Eiger-stimulated apoptosis, respectively, a Tak1 mutant shows dominant genetic interactions with slpr, suggesting potential redundant or combinatorial functions. Finally, we demonstrate that SLPR overexpression can induce ectopic JNK signaling and that the SLPR protein is enriched at the epithelial cell cortex.     

Sex determination in the germ line of Drosophila depends on genetic signals and inductive somatic factors

We have analyzed the mechanism of sex determination in the germ line of Drosophila by manipulating three parameters: (1) the ratio of X-chromosomes to sets of autosomes (X:A); (2) the state of activity of the gene Sex-lethal (Sxl), and (3) the sex of the gonadal soma. To this end, animals with a ratio of 2X:2A and 2X:3A were sexually transformed into pseudomales by mutations at the sex-determining genes Sxl (Sex-lethal), tra (transformer), tra-2 (transformer-2), or dsx (double-sex). Animals with the karyotype 2X;3A were also transformed into pseudofemales by the constitutive mutation SxlM1. The sexual phenotype of the gonads and of the germ cells was assessed by phase-contrast microscopy. Confirming the conclusions of Steinmann-Zwicky et al. (Cell 57, 157, 1989), we found that all three parameters affect sex determination in germ cells. In contrast to the soma in which sex determination is completely cell-autonomous, sex determination in the germ line has a non-autonomous component inasmuch as the sex of the soma can influence the sexual pathway of the germ cells. Somatic induction has a clear effect on 2X;2A germ cells that carry a Sxl+ allele. These cells, which form eggs in an ovary, can enter spermatogenesis in testes. Mutations that cause partial loss of function or gain of function of Sxl thwart somatic induction and, independently of the sex of the soma, dictate spermatogenesis or oogenesis, respectively. Somatic induction has a much weaker effect on 2X;3A germ cells. This ratio is essentially a male signal for germ cells which consistently enter spermatogenesis in testes, even when they carry SxlM1. In a female soma, however, SxlM1 enables the 2X;3A germ cells to form almost normal eggs. Our results show that sex determination in the germ line is more complex than in the soma. They provide further evidence that the state of Sxl, the key gene for sex determination and dosage compensation in the soma, also determines the sex of the germ cells, and that, in the germ line, the state of activity of Sxl is regulated not only by the X:A ratio, but also by somatic inductive stimuli.