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Jeffrey C Hall

Publications and source records attributed to Jeffrey C Hall.

At least 19 recordsLinked to original sources
PubMed2006

Neurotoxic protein expression reveals connections between the circadian clock and mating behavior in Drosophila.

To investigate the functions of circadian neurons, we added two strategies to the standard Drosophila behavioral genetics repertoire. The first was to express a polyglutamine-expanded neurotoxic protein (MJDtr78Q; MJD, Machado-Joseph disease) in the major timeless (tim)-expressing cells of the adult brain. These Tim-MJD flies were viable, in contrast to the use of cell-death gene expression for tim neuron inactivation. Moreover, they were more arrhythmic than flies expressing other neurotoxins and had low but detectable tim mRNA levels. The second extended standard microarray technology from fly heads to dissected fly brains. By combining the two approaches, we identified a population of Tim-MJD-affected mRNAs. Some had been previously identified as sex-specific and relevant to courtship, including mRNAs localized to brain-proximal fat-body tissue and brain courtship centers. Finally, we found a decrease in the number of neurons that expressed male-specific forms of the fruitless protein in the laterodorsal region of the brain. The decrease was not a consequence of toxic protein expression within these specialized cells but a likely effect of communication with neighboring TIM-expressing neurons. The data suggest a functional interaction between adjacent circadian and mating circuits within the fly brain, as well as an interaction between circadian circuits and brain-proximal fat body.

Animals↗
PubMed2006

Defective transfer of seminal-fluid materials during matings of semi-fertile fruitless mutants in Drosophila.

In context of the semi-sterility exhibited by Drosophila males expressing certain mating-enabling fruitless (fru) mutant genotypes, we examined the transfer of seminal fluid using a transgene that encodes the Sex Peptide (SP) oligopeptide fused to Green Fluorescent Protein (GFP). We found that this fusion construct expresses SP-GFP in a valid manner within accessory glands of the male reproductive system in normal and fru-mutant males. Transfer of SP-GFP to live females was readily detectable during and after copulation. With respect to the pertinent combinations of fru mutations, we demonstrated that these abnormal genotypes cause males to transmit mating-related materials in two aberrant ways: one involving whether any seminal-fluid entities are transferred at all during a given mating; the other revealing an intriguing aspect of these fruitless effects, such that the mutations in question cause males to transfer female-affecting materials in a manner that varies among copulations. In this regard, certain mutant males that do not transfer SP nevertheless are able to transfer sperm: a fru-mated female possessing no GFP who was not fecund initially could produce progeny when seminal-fluid proteins were subsequently supplied by mating with a male that was spermless owing to the effects of a tudor mutation.

Analysis of Variance↗
PubMed2005

Functional analysis of fruitless gene expression by transgenic manipulations of Drosophila courtship.

A gal4-containing enhancer-trap called C309 was previously shown to cause subnormal courtship of Drosophila males toward females and courtship among males when driving a conditional disrupter of synaptic transmission (shi(TS)). We extended these manipulations to analyze all features of male-specific behavior, including courtship song, which was almost eliminated by driving shi(TS) at high temperature. In the context of singing defects and homosexual courtship affected by mutations in the fru gene, a tra-regulated component of the sex-determination hierarchy, we found a C309/tra(F) combination also to induce high levels of courtship between pairs of males and "chaining" behavior in groups; however, these doubly transgenic males sang normally. Because production of male-specific FRU(M) protein is regulated by TRA, we hypothesized that a fru-derived transgene encoding the male (M) form of an Inhibitory RNA (fru(MIR)) would mimic the effects of tra(F); but C309/fru(MIR) males exhibited no courtship chaining, although they courted other males in single-pair tests. Double-labeling of neurons in which GFP was driven by C309 revealed that 10 of the 20 CNS clusters containing FRU(M) in wild-type males included coexpressing neurons. Histological analysis of the developing CNS could not rationalize the absence of tra(F) or fru(MIR) effects on courtship song, because we found C309 to be coexpressed with FRU(M) within the same 10 neuronal clusters in pupae. Thus, we hypothesize that elimination of singing behavior by the C309/shi(TS) combination involves neurons acting downstream of FRU(M) cells.

Animals↗
PubMed2005

Male-specific fruitless specifies the neural substrates of Drosophila courtship behaviour.

Robust innate behaviours are attractive systems for genetically dissecting how environmental cues are perceived and integrated to generate complex behaviours. During courtship, Drosophila males engage in a series of innate, stereotyped behaviours that are coordinated by specific sensory cues. However, little is known about the specific neural substrates mediating this complex behavioural programme. Genetic, developmental and behavioural studies have shown that the fruitless (fru) gene encodes a set of male-specific transcription factors (FruM) that act to establish the potential for courtship in Drosophila. FruM proteins are expressed in approximately 2% of central nervous system neurons, at least one subset of which coordinates the component behaviours of courtship. Here we have inserted the yeast GAL4 gene into the fru locus by homologous recombination and show that (1) FruM is expressed in subsets of all peripheral sensory systems previously implicated in courtship, (2) inhibition of FruM function in olfactory system components reduces olfactory-dependent changes in courtship behaviour, (3) transient inactivation of all FruM-expressing neurons abolishes courtship behaviour, with no other gross changes in general behaviour, and (4) 'masculinization' of FruM-expressing neurons in females is largely sufficient to confer male courtship behaviour. Together, these data demonstrate that FruM proteins specify the neural substrates of male courtship.

Animals↗
PubMed2005

Comparative analysis of Corazonin-encoding genes (Crz's) in Drosophila species and functional insights into Crz-expressing neurons.

To gain insight into regulatory mechanisms of tissue-specific Corazonin (Crz) gene expression and its functions in Drosophila, we cloned the Crz genes from four Drosophila species (D. melanogaster, D. simulans, D. erecta, and D. virilis) and performed comparative analyses of Crz gene sequences and expression patterns using in situ hybridization and immunohistochemistry. Although Crz gene sequences showed a great deal of diversity, its expression patterns in the CNS were highly conserved in the Drosophila species examined here. In D. melanogaster larva, Crz expression was found in four pairs of neurons per cerebral lobe and in eight pairs of bilateral neurons in the ventral nerve cord; in adult, the number of Crz-producing neurons increased to 6-8 in the pars lateralis of each brain lobe, whereas neurons in the ventral nerve cord were no longer detectable. Crz transcripts were also found in the optic lobes; however, these mRNAs do not seem to be translated. Such adult-like Crz expression patterns were established within 48 hours after pupation. Somata of Crz-neurons in the pars lateralis are located in the vicinity of terminals emanating from PDF-containing pacemaking neurons, indicating a functional connection between the two peptidergic nervous systems. A subset of Crz neurons coexpressed the period clock gene; however, normal Crz transcription was unaffected by central clockworks. Two pairs of ectopic Crz cells were detected in the adult brains of behaviorally arrhythmic Clock(Jrk) or cycle(02) mutants, suggesting that CLOCK and CYCLE proteins negatively regulate Crz transcription in a cell-specific manner.

Animals↗
PubMed2005

Systems approaches to biological rhythms in Drosophila.

The chronobiological system of Drosophila is considered from the perspective of rhythm-regulated genes. These factors are enumerated and discussed not so much in terms of how the gene products are thought to act on behalf of circadian-clock mechanisms, but with special emphasis on where these molecules are manufactured within the organism. Therefore, with respect to several such cell and tissue types in the fly head, what is the "systems meaning" of a given structure's function insofar as regulation of rest-activity cycles is concerned? (Systematic oscillation of daily behavior is the principal overt phenotype analyzed in studies of Drosophila chronobiology). In turn, how do the several separate sets of clock-gene-expressing cells interact--or in some cases act in parallel--such that intricacies of the fly's sleep-wake cycles are mediated? Studying Drosophila chrono-genetics as a system-based endeavor also encompasses the fact that rhythm-related genes generate their products in many tissues beyond neural ones and during all stages of the life cycle. What, then, is the meaning of these widespread gene-expression patterns? This question is addressed with regard to circadian rhythms outside the behavioral arena, by considering other kinds of temporally based behaviors, and by contemplating how broadly systemic expression of rhythm-related genes connects with even more pleiotropic features of Drosophila biology. Thus, chronobiologically connected factors functioning within this insect comprise an increasingly salient example of gene versatility--multi-faceted usages of, and complex interactions among, entities that set up an organism's overall wherewithal to form and function. A corollary is that studying Drosophila development and adult-fly actions, even when limited to analysis of rhythm-systems phenomena, involves many of the animal's tissues and phenotypic capacities. It follows that such chronobiological experiments are technically demanding, including the necessity for investigators to possess wide-ranging expertise. Therefore, this chapter includes several different kinds of Methods set-asides. These techniques primers necessarily lack comprehensiveness, but they include certain discursive passages about why a given method can or should be applied and concerning real-world applicability of the pertinent rhythm-related technologies.

Animals↗
PubMed2003

A self-sustaining, light-entrainable circadian oscillator in the Drosophila brain.

BACKGROUND: The circadian clock of Drosophila is able to drive behavioral rhythms for many weeks in continuous darkness (DD). The endogenous rhythm generator is thought to be generated by interlocked molecular feedback loops involving circadian transcriptional and posttranscriptional regulation of several clock genes, including period. However, all attempts to demonstrate sustained rhythms of clock gene expression in DD have failed, making it difficult to link the molecular clock models with the circadian behavioral rhythms. Here we restricted expression of a novel period-luciferase transgene to certain clock neurons in the Drosophila brain, permitting us to monitor reporter gene activity in these cells in real-time. RESULTS: We show that only a subset of the previously described pacemaker neurons is able to sustain PERIOD protein oscillations after 5 days in constant darkness. In addition, we identified a sustained and autonomous molecular oscillator in a group of clock neurons in the dorsal brain with heretofore unknown function. We found that these "dorsal neurons" (DNs) can synchronize behavioral rhythms and that light input into these cells involves the blue-light photoreceptor cryptochrome. CONCLUSIONS: Our results suggest that the DNs play a prominent role in controlling locomotor behavior when flies are exposed to natural light-dark cycles. Analysis of similar "stable mosaic" transgenes should help to reveal the function of the other clock neuronal clusters within the fly brain.

Animals↗
PubMed2003

Drosophila free-running rhythms require intercellular communication.

Robust self-sustained oscillations are a ubiquitous characteristic of circadian rhythms. These include Drosophila locomotor activity rhythms, which persist for weeks in constant darkness (DD). Yet the molecular oscillations that underlie circadian rhythms damp rapidly in many Drosophila tissues. Although much progress has been made in understanding the biochemical and cellular basis of circadian rhythms, the mechanisms that underlie the differences between damped and self-sustaining oscillations remain largely unknown. A small cluster of neurons in adult Drosophila brain, the ventral lateral neurons (LN(v)s), is essential for self-sustained behavioral rhythms and has been proposed to be the primary pacemaker for locomotor activity rhythms. With an LN(v)-specific driver, we restricted functional clocks to these neurons and showed that they are not sufficient to drive circadian locomotor activity rhythms. Also contrary to expectation, we found that all brain clock neurons manifest robust circadian oscillations of timeless and cryptochrome RNA for many days in DD. This persistent molecular rhythm requires pigment-dispersing factor (PDF), an LN(v)-specific neuropeptide, because the molecular oscillations are gradually lost when Pdf(01) mutant flies are exposed to free-running conditions. This observation precisely parallels the previously reported effect on behavioral rhythms of the Pdf(01) mutant. PDF is likely to affect some clock neurons directly, since the peptide appears to bind to the surface of many clock neurons, including the LN(v)s themselves. We showed that the brain circadian clock in Drosophila is clearly distinguishable from the eyes and other rapidly damping peripheral tissues, as it sustains robust molecular oscillations in DD. At the same time, different clock neurons are likely to work cooperatively within the brain, because the LN(v)s alone are insufficient to support the circadian program. Based on the damping results with Pdf(01) mutant flies, we propose that LN(v)s, and specifically the PDF neuropeptide that it synthesizes, are important in coordinating a circadian cellular network within the brain. The cooperative function of this network appears to be necessary for maintaining robust molecular oscillations in DD and is the basis of sustained circadian locomotor activity rhythms.

Animals↗
PubMed2003

Genetics and molecular biology of rhythms in Drosophila and other insects.

Application of generic variants (Sections II-IV, VI, and IX) and molecular manipulations of rhythm-related genes (Sections V-X) have been used extensively to investigate features of insect chronobiology that might not have been experimentally accessible otherwise. Most such tests of mutants and molecular-genetic xperiments have been performed in Drosophila melanogaster. Results from applying visual-system variants have revealed that environmental inputs to the circadian clock in adult flies are mediated by external photoreceptive structures (Section II) and also by direct light reception chat occurs in certain brain neurons (Section IX). The relevant light-absorbing molecuLes are rhodopsins and "blue-receptive" cryptochrome (Sections II and IX). Variations in temperature are another clock input (Section IV), as has been analyzed in part by use of molecular techniques and transgenes involving factors functioning near the heart of the circadian clock (Section VIII). At that location within the fly's chronobiological system, approximately a half-dozen-perhaps up to as many as 10-clock genes encode functions that act and interact to form the circadian pacemaker (Sections III and V). This entity functions in part by transcriptional control of certain clock genes' expressions, which result in the production of key proteins that feed back negatively to regulate their own mRNA production. This occurs in part by interactions of such proteins with others that function as transcriptional activators (Section V). The implied feedback loop operates such that there are daily variations in the abundances of products put out by about one-half of the core clock genes. Thus, the normal expression of these genes defines circadian rhythms of their own, paralleling the effects of mutations at the corresponding genetic loci (Section III), which are to disrupt or apparently eliminate clock functioning. The fluctuations in the abundance of gene products are controlled transciptionally and posttranscriptionally. These clock mechanisms are being analyzed in ways that are increasingly complex and occasionally obscure; not all panels of this picture are comprehensive or clear, including problems revolving round the biological meaning or a given features of all this molecular cycling (Section V). Among the complexities and puzzles that have recently arisen, phenomena that stand out are posttranslational modifications of certain proteins that are circadianly regulated and regulating; these biochemical events form an ancillary component of the clock mechanism, as revealed in part by genetic identification of Factors (Section III) that turned out to encode protein kinases whose substrates include other pacemaking polypeptides (Section V). Outputs from insect circadian clocks have been long defined on formalistic and in some cases concrete criteria, related to revealed rhythms such as periodic eclosion and daily fluctuations of locomotion (Sections II and III). Based on the reasoning that if clock genes can regulate circadian cyclings of their own products, they can do the same for genes that function along output pathways; thus clock-regulated genes have been identified in part by virtue of their products' oscillations (Section X). Those studied most intensively have their expression influenced by circadian-pacemaker mutations. The clock-regulated genes discovered on molecular criteria have in some instances been analyzed further in their mutant forms and found to affect certain features of overt whole-organismal rhythmicity (Sections IV and X). Insect chronogenetics touches in part on naturally occurring gene variations that affect biological rhythmicity or (in some cases) have otherwise informed investigators about certain features of the organism's rhythm system (Section VII). Such animals include at least a dozen insect species other than D. melanogaster in which rhythm variants have been encountered (although usually not looked for systematically). The chronobiological "system" in the fruit fly might better be graced with a plural appellation because there is a myriad of temporally related phenomena that have come under the sway of one kind of putative rhythm variant or the other (Section IV). These phenotypes, which range well beyond the bedrock eclosion and locomotor circadian rhythms, unfortunately lead to the creation of a laundry list of underanalyzed or occult phenomena that may or may not be inherently real, whether or not they might be meaningfully defective under the influence of a given chronogenetic variant. However, such mutants seem to lend themselves to the interrogation of a wide variety of time-based attributes-those that fall within the experimental confines of conventionally appreciated circadian rhythms (Sections II, III, VI, and X); and others that consist of 24-hr or nondaily cycles defined by many kinds of biological, physiological, or biochemical parameters (Section IV).

Animals↗
PubMed2002

Resetting the circadian clock by social experience in Drosophila melanogaster.

Circadian clocks are influenced by social interactions in a variety of species, but little is known about the sensory mechanisms underlying these effects. We investigated whether social cues could reset circadian rhythms in Drosophila melanogaster by addressing two questions: Is there a social influence on circadian timing? If so, then how is that influence communicated? The experiments show that in a social context Drosophila transmit and receive cues that influence circadian time and that these cues are likely olfactory.

Animals↗
PubMed2002

Advanced analysis of a cryptochrome mutation's effects on the robustness and phase of molecular cycles in isolated peripheral tissues of Drosophila.

BACKGROUND: Previously, we reported effects of the cry(b) mutation on circadian rhythms in period and timeless gene expression within isolated peripheral Drosophila tissues. We relied on luciferase activity driven by the respective regulatory genomic elements to provide real-time reporting of cycling gene expression. Subsequently, we developed a tool kit for the analysis of behavioral and molecular cycles. Here, we use these tools to analyze our earlier results as well as additional data obtained using the same experimental designs. RESULTS: Isolated antennal pairs, heads, bodies, wings and forelegs were evaluated under light-dark cycles. In these conditions, the cry(b) mutation significantly decreases the number of rhythmic specimens in each case except the wing. Moreover, among those specimens with detectable rhythmicity, mutant rhythms are significantly weaker than cry+ controls. In addition, cry(b) alters the phase of period gene expression in these tissues. Furthermore, peak phase of luciferase-reported period and timeless expression within cry+ samples is indistinguishable in some tissues, yet significantly different in others. We also analyze rhythms produced by antennal pairs in constant conditions. CONCLUSIONS: These analyses further show that circadian clock mechanisms in Drosophila may vary in a tissue-specific manner, including how the cry gene regulates circadian gene expression.

5' Flanking Region↗
PubMed2002

Signal analysis of behavioral and molecular cycles.

BACKGROUND: Circadian clocks are biological oscillators that regulate molecular, physiological, and behavioral rhythms in a wide variety of organisms. While behavioral rhythms are typically monitored over many cycles, a similar approach to molecular rhythms was not possible until recently; the advent of real-time analysis using transgenic reporters now permits the observations of molecular rhythms over many cycles as well. This development suggests that new details about the relationship between molecular and behavioral rhythms may be revealed. Even so, behavioral and molecular rhythmicity have been analyzed using different methods, making such comparisons difficult to achieve. To address this shortcoming, among others, we developed a set of integrated analytical tools to unify the analysis of biological rhythms across modalities. RESULTS: We demonstrate an adaptation of digital signal analysis that allows similar treatment of both behavioral and molecular data from our studies of Drosophila. For both types of data, we apply digital filters to extract and clarify details of interest; we employ methods of autocorrelation and spectral analysis to assess rhythmicity and estimate the period; we evaluate phase shifts using crosscorrelation; and we use circular statistics to extract information about phase. CONCLUSION: Using data generated by our investigation of rhythms in Drosophila we demonstrate how a unique aggregation of analytical tools may be used to analyze and compare behavioral and molecular rhythms. These methods are shown to be versatile and will also be adaptable to further experiments, owing in part to the non-proprietary nature of the code we have developed.

Activity Cycles↗
PubMed2002

Identification of circadian-clock-regulated enhancers and genes of Drosophila melanogaster by transposon mobilization and luciferase reporting of cyclical gene expression.

A new way was developed to isolate rhythmically expressed genes in Drosophila by modifying the classic enhancer-trap method. We constructed a P element containing sequences that encode firefly luciferase as a reporter for oscillating gene expression in live flies. After generation of 1176 autosomal insertion lines, bioluminescence screening revealed rhythmic reporter-gene activity in 6% of these strains. Rhythmically fluctuating reporter levels were shown to be altered by clock mutations in genes that specify various circadian transcription factors or repressors. Intriguingly, rhythmic luminescence in certain lines was affected by only a subset of the pacemaker mutations. By isolating genes near 13 of the transposon insertions and determining their temporal mRNA expression pattern, we found that four of the loci adjacent to the trapped enhancers are rhythmically expressed. Therefore, this approach is suitable for identifying genetic loci regulated by the circadian clock. One transposon insert caused a mutation in the rhythmically expressed gene numb. This novel numb allele, as well as previously described ones, was shown to affect the fly's rhythm of locomotor activity. In addition to its known role in cell fate determination, this gene and the phosphotyrosine-binding protein it encodes are likely to function in the circadian system.

Animals↗
PubMed2002

Courtship and other behaviors affected by a heat-sensitive, molecularly novel mutation in the cacophony calcium-channel gene of Drosophila.

The cacophony (cac) locus of Drosophila melanogaster, which encodes a calcium-channel subunit, has been mutated to cause courtship-song defects or abnormal responses to visual stimuli. However, the most recently isolated cac mutant was identified as an enhancer of a comatose mutation's effects on general locomotion. We analyzed the cac(TS2) mutation in terms of its intragenic molecular change and its effects on behaviors more complex than the fly's elementary ability to move. The molecular etiology of this mutation is a nucleotide substitution that causes a proline-to-serine change in a region of the polypeptide near its EF hand. Given that this motif is involved in channel inactivation, it was intriguing that cac(TS2) males generate song pulses containing larger-than-normal numbers of cycles--provided that such males are exposed to an elevated temperature. Similar treatments caused only mild visual-response abnormalities and generic locomotor sluggishness. These results are discussed in the context of calcium-channel functions that subserve certain behaviors and of defects exhibited by the original cacophony mutant. Despite its different kind of amino-acid substitution, compared with that of cac(TS2), cac(S) males sing abnormally in a manner that mimics the new mutant's heat-sensitive song anomaly.

Amino Acid Sequence↗
PubMed2002

Mapping of elements involved in regulating normal temporal period and timeless RNA expression patterns in Drosophila melanogaster.

Although transcriptional regulation is a major force in generating circadian oscillations of clock molecules, posttranscriptional mechanisms also contribute to molecular rhythms. Applying novel transgenic period-luciferase constructs in transgenic Drosophila, the authors show that sequences within per's 5'-untranslated region mediate posttranscriptional regulation at the RNA level. Further mapping suggests that the relevant sequences for the correct phasing of period mRNA expression are located within the first intron. The results are consistent with a clock-regulated temporal stabilization of period mRNA during its daily upswing in the morning. This process is inferred to depend on a function of the PERIOD and TIMELESS proteins, and could further contribute to the observed delay between RNA and protein accumulation. Similarly, applying timeless-luciferase constructs led to the demonstration that regulatory elements for proper temporal timeless expression are present in a 4 kb promoter fragment and in sequences within the first intron. The results establish that, for normal rhythmicity, expression of clock genes requires regulation at the transcriptional, posttranscriptional, and posttranslational levels.

5' Untranslated Regions↗
PubMedDate not supplied

Doublesex gene expression in the central nervous system of Drosophila melanogaster.

Despite several behavior-genetic studies that have suggested roles played by doublesex (dsx) in neural tissues, it has not been demonstrated that the products of this gene are actually present in the central nervous system (CNS). In this report, we describe the cellular, spatial, and temporal expression patterns of dsx gene products in the developing and adult CNS by applying RT-PCR and immunohistochemical procedures. dsx gene products were detected in the CNS of 3rd-instar larvae, pupae, and adults. DSX-immunoreactive signals were observed within the brain and in both the thoracic plus abdominal ganglia of the ventral nerve cord. Most, but not all, cells inferred to contain DSX proteins (by the results of genetic controls for antibody specificity) were further determined to be neurons (by coexpression of a protein that marks such CNS cell types). Temporally varying expression of DSX was most prominently observed in the rapidly metamorphosing early and mid-pupal stages, suggesting that this gene contributes to establishment of sexually dimorphic neuronal structures which subserve adult sexual behaviors. Elements of the spatial and temporal patterns of DSX immunoreactivity also imply that sexually dimorphic dsx expression in certain neuronal clusters within the adult CNS could participate in ongoing operations of the mature nervous system with respect to the courtship behaviors that are affected by dsx mutations.

Animals↗