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M W Young

Publications and source records attributed to M W Young.

At least 37 records · Page 2Linked to original sources
PubMed1999

Molecular control of circadian behavioral rhythms.

Several genes have been recognized in Drosophila that regulate circadian rhythms. Homologues of these genes have now been found in mice and humans, suggesting a mechanism that is conserved throughout the animal kingdom. For some of these genes and their products, molecular oscillations are produced in certain cells of the Drosophila and mammalian brain. Two genes, period and timeless, are transcribed with a circadian rhythm that is regulated by activities derived from their encoded proteins, PER and TIM. Nuclear localization of these proteins downregulates per and tim transcription by suppressing the activities of two transcription factors, dCLOCK and dBMAL1. Cycles in this feedback regulation are promoted by events that regulate the accumulation, physical interaction, and nuclear translocation of PER and TIM proteins. PER and TIM must physically associate to enter the nucleus and their cytoplasmic interaction is delayed by a kinase encoded by the clock gene, double-time. This kinase directs PER phosphorylation, which leads to PER degradation. Effects of the kinase are blocked once PER is complexed to TIM. These interactions prolong the interval of per and tim transcription by ensuring that PER/TIM complexes from only after TIM has accumulated for several hours.

Animals↗
PubMed1998

Ligand-induced cleavage and regulation of nuclear entry of Notch in Drosophila melanogaster embryos.

Notch, a transmembrane protein found in a wide range of organisms, is a component of a pathway that mediates cell-fate decisions that involve intercellular communication. In this paper, we show that in Drosophila melanogaster, Notch (N) is processed in a ligand-dependent fashion to generate phosphorylated, soluble intracellular derivatives. Suppressor of Hairless [Su(H)] is predominantly associated with soluble intracellular N. It has been demonstrated by others that N has access to the nucleus, and we show that when tethered directly to DNA, the cytoplasmic domain of N can activate transcription. Conversely, a viral activator fused to Su(H) can substitute for at least some N functions during embryogenesis. We suggest that one function of soluble forms of N is to bind to Su(H), and in the nucleus, to act directly as a transcriptional transactivator of the latter protein. Although N has functional nuclear localization signals, the N/Su(H) complex accumulates in the cytoplasm and on membranes suggesting that its nuclear entry is regulated. Localization studies in cultured cells and embryos suggest that Su(H) plays a role in this regulation, with the relative levels of Delta, N and Su(H) determining whether a N/Su(H) complex enters the nucleus.

Animals↗
PubMed1998

A light-independent oscillatory gene mPer3 in mouse SCN and OVLT.

A new member of the mammalian period gene family, mPer3, was isolated and its expression pattern characterized in the mouse brain. Like mPer1, mPer2 and Drosophila period, mPer3 has a dimerization PAS domain and a cytoplasmic localization domain. mPer3 transcripts showed a clear circadian rhythm in the suprachiasmatic nucleus (SCN). Expression of mPer3 was not induced by exposure to light at any phase of the clock, distinguishing this gene from mPer1 and mPer2. Cycling expression of mPer3 was also found outside the SCN in the organum vasculosum lamina terminalis (OVLT), a potentially key region regulating rhythmic gonadotropin production and pyrogen-induced febrile phenomena. Thus, mPer3 may contribute to pacemaker functions both inside and outside the SCN.

Amino Acid Sequence↗
PubMed1998

double-time is a novel Drosophila clock gene that regulates PERIOD protein accumulation.

We have isolated three alleles of a novel Drosophila clock gene, double-time (dbt). Short- (dbtS) and long-period (dbtL) mutants alter both behavioral rhythmicity and molecular oscillations from previously identified clock genes, period and timeless. A third allele, dbtP, causes pupal lethality and eliminates circadian cycling of per and tim gene products in larvae. In dbtP mutants, PER proteins constitutively accumulate, remain hypophosphorylated, and no longer depend on TIM proteins for their accumulation. We propose that the normal function of DOUBLETIME protein is to reduce the stability and thus the level of accumulation of monomeric PER proteins. This would promote a delay between per/tim transcription and PER/TIM complex function, which is essential for molecular rhythmicity.

Alleles↗
PubMed1998

The Drosophila clock gene double-time encodes a protein closely related to human casein kinase Iepsilon.

The cloning of double-time (dbt) is reported. DOUBLETIME protein (DBT) is most closely related to human casein kinase Iepsilon. dbtS and dbtL mutations, which alter period length of Drosophila circadian rhythms, produce single amino acid changes in conserved regions of the predicted kinase. dbtP mutants, which eliminate rhythms of per and tim expression and constitutively overproduce hypophosphorylated PER proteins, abolish most dbt expression. dbt mRNA appears to be expressed in the same cell types as are per and tim and shows no evident oscillation in wild-type heads. DBT is capable of binding to PER in vitro and in Drosophila cells, suggesting that a physical association of PER and DBT regulates PER phosphorylation and accumulation in vivo.

Amino Acid Sequence↗
PubMed1998

Mammalian circadian autoregulatory loop: a timeless ortholog and mPer1 interact and negatively regulate CLOCK-BMAL1-induced transcription.

We report the cloning and mapping of mouse (mTim) and human (hTIM) orthologs of the Drosophila timeless (dtim) gene. The mammalian Tim genes are widely expressed in a variety of tissues; however, unlike Drosophila, mTim mRNA levels do not oscillate in the suprachiasmatic nucleus (SCN) or retina. Importantly, hTIM interacts with the Drosophila PERIOD (dPER) protein as well as the mouse PER1 and PER2 proteins in vitro. In Drosophila (S2) cells, hTIM and dPER interact and translocate into the nucleus. Finally, hTIM and mPER1 specifically inhibit CLOCK-BMAL1-induced transactivation of the mPer1 promoter. Taken together, these results demonstrate that mTim and hTIM are mammalian orthologs of timeless and provide a framework for a basic circadian autoregulatory loop in mammals.

ARNTL Transcription Factors↗
PubMed1998

The molecular control of circadian behavioral rhythms and their entrainment in Drosophila.

Molecular and genetic characterizations of circadian rhythms in Drosophila indicate that function of an intracellular pacemaker requires the activities of proteins encoded by three genes: period (per), timeless (tim), and doubletime (dbt). RNA from two of these genes, per and tim, is expressed with a circadian rhythm. Heterodimerization of PER and TIM proteins allows nuclear localization and suppression of further RNA synthesis by a PER/TIM complex. These protein interactions promote cyclical gene expression because heterodimers are observed only at high concentrations of per and tim RNA, separating intervals of RNA accumulation from times of PER/TIM complex activity. Light resets these molecular cycles by eliminating TIM. The product of dbt also regulates accumulation of per and tim RNA, and it may influence action of the PER/TIM complex. The recent discovery of PER homologues in mice and humans suggests that a related mechanism controls mammalian circadian behavioral rhythms.

Animals↗
PubMed1997

Comparison of chromosomal DNA composing timeless in Drosophila melanogaster and D. virilis suggests a new conserved structure for the TIMELESS protein.

Two proteins, TIM and PER, physically interact to control circadian cycles of tim and per transcription in Drosophila melanogaster. In the present study the structure of TIM protein expressed by D. virilis was determined by isolation and sequence analysis of genomic DNA (gDNA) corresponding to the D. virilis tim locus (v tim ). Comparison of v tim and m tim gDNA revealed high conservation of the TIM protein. This contrasts with poor sequence conservation previously observed for the TIM partner protein PER in these species. Inspection of the v tim sequence suggests an alternative structure for most TIM proteins. Sequences forming an intron in a previously characterized D. melanogaster tim cDNA appear to be most often translated to produce a longer TIM protein in both species. The N-terminal sequence of vTIM and sequence analysis of genomic DNA from several strains of D. melanogaster suggest that only one of two possible translation initiation sites found in tim mRNA is sufficient to generate circadian rhythms in D. melanogaster. TIM translation may be affected by multiple AUG codons that appear to have been conserved in sequences composing the 5'-untranslated tim mRNA leader.

Amino Acid Sequence↗
PubMed1997

Diverse roles for the Notch receptor in the development of D. melanogaster.

Notch proteins appear to be involved in cell fate commitments with deep evolutionary roots. Homologues have been shown to play key roles in the development of nematodes, insects, amphibia, and mammals. Activity of the Notch receptor has been observed in the patterning of ectoderm, mesoderm, and endoderm, indicating an origin prior to the functional differentiation of these germ layers. To understand how a single receptor can participate so widely in development, we have been examining the role of specific extracellular segments of Notch. Early studies of mutations affecting widely separated EGF-like elements of Notch first raised the possibility for interaction with multiple ligands. Biochemical approaches, and exhaustive structure function studies in transgenic Drosophila are beginning to reveal how this receptor is activated, and point to a range of physical interactions with other proteins.

Animals↗
PubMed1996

Light-induced degradation of TIMELESS and entrainment of the Drosophila circadian clock.

Two genes, period (per) and timeless (tim), are required for production of circadian rhythms in Drosophila. The proteins encoded by these genes (PER and TIM) physically interact, and the timing of their association and nuclear localization is believed to promote cycles of per and tim transcription through an autoregulatory feedback loop. Here it is shown that TIM protein may also couple this molecular pacemaker to the environment, because TIM is rapidly degraded after exposure to light. TIM accumulated rhythmically in nuclei of eyes and in pacemaker cells of the brain. The phase of these rhythms was differentially advanced or delayed by light pulses delivered at different times of day, corresponding with phase shifts induced in the behavioral rhythms.

Animals↗
PubMed1996

Regulation of nuclear entry of the Drosophila clock proteins period and timeless.

Two genes, period (per) and timeless (tim), are essential for circadian rhythmicity in Drosophila. The encoded proteins (PER and TIM) physically interact. Here, it is shown that TIM and PER accumulate in the cytoplasm when independently expressed in cultured (S2) Drosophila cells. However, the proteins move to the nuclei of these cells if coexpressed. Domains of PER and TIM have been identified that block nuclear localization of the monomeric proteins. In vitro protein interaction studies indicate that the sequence inhibiting the nuclear accumulation of PER forms a binding site for TIM. The results indicate a mechanism for controlled nuclear localization in which suppression of cytoplasmic localization is accomplished by direct interaction of PER and TIM. No other clock functions are required for nuclear localization. The findings suggest that a checkpoint in the circadian cycle is established by requiring cytoplasmic assembly of a PER/TIM complex as a condition for nuclear transport of either protein.

Animals↗
PubMed1995

Positional cloning and sequence analysis of the Drosophila clock gene, timeless.

The Drosophila genes timeless (tim) and period (per) interact, and both are required for production of circadian rhythms. Here the positional cloning and sequencing of tim are reported. The tim gene encodes a previously uncharacterized protein of 1389 amino acids, and possibly another protein of 1122 amino acids. The arrhythmic mutation tim01 is a 64-base pair deletion that truncates TIM to 749 amino acids. Absence of sequence similarity to the PER dimerization motif (PAS) indicates that direct interaction between PER and TIM would require a heterotypic protein association.

Amino Acid Sequence↗
PubMed1995

Rhythmic expression of timeless: a basis for promoting circadian cycles in period gene autoregulation.

The clock gene timeless (tim) is required for circadian rhythmicity in Drosophila. The accumulation of tim RNA followed a circadian rhythm, and the phase and period of the tim RNA rhythm were indistinguishable from those that have been reported for per. The tim RNA oscillations were found to be dependent on the presence of PER and TIM proteins, which demonstrates feedback control of tim by a mechanism previously shown to regulate per expression. The cyclic expression of tim appears to dictate the timing of PER protein accumulation and nuclear localization, suggesting that tim promotes circadian rhythms of per and tim transcription by restricting per RNA and PER protein accumulation to separate times of day.

Animals↗
PubMed1995

Isolation of timeless by PER protein interaction: defective interaction between timeless protein and long-period mutant PERL.

The period (per) gene likely encodes a component of the Drosophila circadian clock. Circadian oscillations in the abundance of per messenger RNA and per protein (PER) are thought to arise from negative feedback control of per gene transcription by PER. A recently identified second clock locus, timeless (tim), apparently regulates entry of PER into the nucleus. Reported here are the cloning of complementary DNAs derived from the tim gene in a two-hybrid screen for PER-interacting proteins and the demonstration of a physical interaction between the tim protein (TIM) and PER in vitro. A restricted segment of TIM binds directly to a part of the PER dimerization domain PAS. PERL, a mutation that causes a temperature-sensitive lengthening of circadian period and a temperature-sensitive delay in PER nuclear entry, exhibits a temperature-sensitive defect in binding to TIM. These results suggest that the interaction between TIM and PER determines the timing of PER nuclear entry and therefore the duration of part of the circadian cycle.

Animals↗
PubMed1995

Suppression of PERIOD protein abundance and circadian cycling by the Drosophila clock mutation timeless.

The timeless mutation (tim) leads to loss of circadian behavioral rhythms in Drosophila melanogaster. The effects of tim on rhythmicity involve interactions with period (per), a second essential clock gene, as the tim mutation suppresses circadian oscillations of per transcription and blocks nuclear localization of a PER reporter protein. In the present study it was found that the tim mutant constitutively produces a low level of PER protein that is comparable with that produced late in the day by wild-type flies. In addition, it was shown that tim suppresses circadian cycling of PER protein abundance and circadian regulation of PER phosphorylation. Transfer of wild-type flies to constant light also suppressed cycling of PER abundance and phosphorylation and produced constitutively low levels of PER. In the tim mutant there was no additional effect of constant light on PER. These results suggest that constant light and the tim mutation produce related changes in the underlying biological clock. We further suggest that the multiple effects of tim are due to a primary effect on per expression at the posttranscriptional level. The effects of tim on behavioral rhythms and per RNA cycling are therefore likely to involve effects on PER protein through previously proposed feedback mechanisms.

Animals↗
PubMed1995

Circadian rhythms in Drosophila can be driven by period expression in a restricted group of central brain cells.

Neural tissues controlling circadian rhythmicity have been identified in a variety of organisms and are often closely associated with the visual system. In Drosophila, the clock gene period (per), which is required for circadian rhythms, is expressed in many neurons and glia throughout the eye and brain. We asked whether biological rhythms could be generated if per expression were restricted to a subset of these cells that is involved in photoreception. Here we demonstrate that expression of per under the control of the glass promoter confers both behavioral and molecular rhythmicity. glass is required for development of Drosophila photoreceptors, and this promoter is active in eyes, ocelli, and certain cells of the central brain. When we genetically removed all external photoreceptor cells, rhythms persisted in these transgenic animals. This suggests that a few central brain cells producing glass and per are capable of generating biological rhythms.

Animals↗