Search PubMed⌕ Search

Biomedical subjects

D Whitmore

Publications and source records attributed to D Whitmore.

18 recordsLinked to original sources
PubMed2000

Asynchronous oscillations of two zebrafish CLOCK partners reveal differential clock control and function.

Most clock genes encode transcription factors that interact to elicit cooperative control of clock function. Using a two-hybrid system approach, we have isolated two different partners of zebrafish (zf) CLOCK, which are similar to the mammalian BMAL1 (brain and muscle arylhydrocarbon receptor nuclear translocator-like protein 1). The two homologs, zfBMAL1 and zfBMAL2, contain conserved basic helix-loop-helix-PAS (Period-Arylhydrocarbon receptor-Singleminded) domains but diverge in the carboxyl termini, thus bearing different transcriptional activation potential. As for zfClock, the expression of both zfBmals oscillates in most tissues in the animal. However, in many tissues, the peak, levels, and kinetics of expression are different between the two genes and for the same gene from tissue to tissue. These results support the existence of independent peripheral oscillators and suggest that zfBMAL1 and zfBMAL2 may exert distinct circadian functions, interacting differentially with zfCLOCK at various times in different tissues. Our findings also indicate that multiple controls may be exerted by the central clock and/or that peripheral oscillators can differentially interpret central clock signals.

ARNTL Transcription Factors↗
PubMed2000

Light acts directly on organs and cells in culture to set the vertebrate circadian clock.

The expression of clock genes in vertebrates is widespread and not restricted to classical clock structures. The expression of the Clock gene in zebrafish shows a strong circadian oscillation in many tissues in vivo and in culture, showing that endogenous oscillators exist in peripheral organs. A defining feature of circadian clocks is that they can be set or entrained to local time, usually by the environmental light-dark cycle. An important question is whether peripheral oscillators are entrained to local time by signals from central pacemakers such as the eyes or are themselves directly light-responsive. Here we show that the peripheral organ clocks of zebrafish are set by light-dark cycles in culture. We also show that a zebrafish-derived cell line contains a circadian oscillator, which is also directly light entrained.

Animals↗
PubMed2000

Rhythmic transcription: the molecular basis of oscillatory melatonin synthesis.

Pulsatile hormone synthesis and secretion are characteristic features of various oscillatory biological systems. Circadian rhythms are critical in the regulation of most physiological functions, and much interest has been centred on the understanding of the molecular mechanisms governing them. Adaptation to a changing environment is an essential feature of physiological regulation. The day-night rhythm is translated into hormonal oscillations governing the metabolism of all living organisms. In mammals the pineal gland is responsible for the circadian synthesis of the hormone melatonin in response to signals originating from the endogenous clock located in the hypothalamic suprachiasmatic nucleus (SCN). The molecular mechanisms involved in rhythmic synthesis of melatonin involve the CREM gene, which encodes transcription factors responsive to activation of the cAMP signalling pathway. The CREM product, ICER, is rhythmically expressed and participates in a transcriptional autoregulatory loop which also controls the amplitude of oscillations of serotonin N-acetyl transferase, the rate-limiting enzyme of melatonin synthesis. Thus, a transcription factor modulates the oscillatory levels of a hormone.

Animals↗
PubMed1998

PASting together the mammalian clock.

Over the past year, the first components of the mammalian clock have been identified; Clock, bmal1 and three homologs of Drosophila period have been cloned, all of which encode PAS proteins. Expression of the mammalian period gene oscillates in many tissues in vivo and in immortalized cell cultures in vitro. Now, can we say that every cell has a circadian clock?

ARNTL Transcription Factors↗
PubMed1998

Zebrafish Clock rhythmic expression reveals independent peripheral circadian oscillators.

The only vertebrate clock gene identified by mutagenesis is mouse Clock, which encodes a bHLH-PAS transcription factor. We have cloned Clock in zebrafish and show that, in contrast to its mouse homologue, it is expressed with a pronounced circadian rhythm in the brain and in two defined pacemaker structures, the eye and the pineal gland. Clock oscillation was also found in other tissues, including kidney and heart. In these tissues, expression of Clock continues to oscillate in vitro. This demonstrates that self-sustaining circadian oscillators exist in several vertebrate organs, as was previously reported for invertebrates.

Amino Acid Sequence↗
PubMed1997

Rhythmic transcription: the molecular basis of circadian melatonin synthesis.

Adaptation to a changing environment is an essential feature of physiological regulation. The day/night rhythm is translated into hormonal oscillations governing the physiology of all living organisms. In mammals the pineal gland is responsible for the synthesis of the hormone melatonin in response to signals originating from the endogenous clock located in the hypothalamic suprachiasmatic nucleus (SCN). The molecular mechanisms involved in rhythmic synthesis of melatonin involve the CREM gene, which encodes transcription factors responsive to activation of the cAMP signalling pathway. The CREM product, ICER, is rhythmically expressed and participates in a transcriptional autoregulatory loop which also controls the amplitude of oscillations of serotonin N-acetyl transferase (AANAT), the rate-limiting enzyme of melatonin synthesis. In contrast, chick pinealocytes possess an endogenous circadian pacemaker which directs AANAT rhythmic expression. cAMP-responsive activator transcription factors CREB and ATF1 and the repressor ICER are highly conserved in the chick with the notable exception of ATF1 that possesses two glutamine-rich domains in contrast to the single domain encountered to date in mammalian systems. ICER is cAMP inducible and undergoes a characteristic day-night oscillation in expression reminiscent of AA-NAT, but with a peak towards the end of the night. Interestingly CREB appears to be phosphorylated constitutively with a transient fall occurring at the beginning of the night. Thus, a transcription factor modulates the oscillatory levels of a hormone.

Acetylserotonin O-Methyltransferase↗
PubMed1996

Circadian rhythm generation, expression and entrainment in a molluscan model system.

The Bulla ocular pacemaker provides remarkable opportunities for cellular study of circadian pacemaker systems. The demonstration of circadian oscillations within individual neurons maintained in culture provides us with a first occasion to study the biophysical and biochemical properties of bona fide neuronal circadian pacemakers. The ocular clock is robust and shares formal similarity with other circadian systems. The development of molecular techniques that can be applied to single neurons should allow research on the Bulla retina to continue to progress towards a molecular analysis of circadian timekeeping.

Animals↗
PubMed1996

Evidence for a central role of transcription in the timing mechanism of a circadian clock.

The retinal circadian clock in the isolated in vitro eye of the marine mollusc Bulla gouldiana exhibits a phase-dependent requirement for transcription. The transcription-sensitive phase extends through most of the subjective day and therefore is substantially longer than the previously reported translation-sensitive phase. Lower concentrations of transcription inhibitors yield a significant dose-dependent lengthening of circadian period. Clock motion can be stopped by a high concentration of the transcription inhibitor 5,6-dichlorobenz-imidazole riboside (DRB) when applied during the sensitive phase; after withdrawal of the inhibitor, motion resumes from the phase at which it was stopped. In a double-pulse experiment, phase shifts to light pulses applied after DRB pulses, and not during the translation-sensitive phase, indicate that the inhibition of transcription has immediate effects on the phase of the clock. These data suggest that DRB-induced phase shifts are independent of translation, which implies that the rate of transcription itself plays a significant role in the mechanism underlying the generation of the circadian cycle.

Animals↗
PubMed1995

Cellular analysis of a molluscan retinal biological clock.

The eye of the opisthobranch mollusc Bulla gouldiana expresses a circadian rhythm in optic nerve impulse frequency. The circadian rhythm is generated among approximately 100 neurons at the base of the retina referred to as basal retinal neurons. These cells are electrically coupled to one another and fire spontaneous action potentials in synchrony. Basal retinal neurons recorded intracellularly exhibit a circadian rhythm in membrane potential that appears to be driven by a circadian modulation of membrane conductance. Membrane conductance is relatively high during the subjective night and decreases after subjective dawn. Recent experiments in our laboratory indicate that individual basal retinal neurons in culture can express circadian rhythms in membrane conductance. When completely isolated, these cells continue to show circadian conductance changes. These studies provide the first direct demonstration that individual neurons can act as circadian pacemakers. Although the precise details of the mechanism generating the circadian periodicity remain obscure, our research indicates that several transmembrane ionic fluxes are not involved in rhythm generation, but that a transmembrane Ca2+ flux is critical for entrainment. Both transcription and translation appear to play critical roles in generating the circadian cycle.

Animals↗
PubMed1994

Calcium plays a central role in phase shifting the ocular circadian pacemaker of Aplysia.

The eye of the marine mollusk Aplysia californica contains an oscillator that drives a circadian rhythm of spontaneous compound action potentials in the optic nerve. Both light and serotonin are known to influence the phase of this ocular rhythm. The aim of the present study was to evaluate the role of extracellular calcium in both light and serotonin-mediated phase shifts. Low calcium treatments were found to cause phase shifts which resembled those produced by the transmitter serotonin. However, unlike serotonin, low calcium neither increased ocular cAMP levels nor could these phase shifts be prevented by increasing extracellular potassium concentration. Low calcium-induced phase shifts were prevented by the simultaneous application of the translational inhibitor anisomycin and low calcium treatment resulted in changes in [35S]methionine incorporation into several proteins as measured by a two-dimensional electrophoresis gel analysis. Finally, light treatments failed to produce phase shifts in the presence of low calcium or the calcium channel antagonist nickel chloride. These results are consistent with a model in which serotonin phase shifts the ocular pacemaker by decreasing a transmembrane calcium flux through membrane hyperpolarization while light-induced phase shifts are mediated by an increase in calcium flux.

Action Potentials↗
PubMed1994

Tourette's syndrome in a special education population: a pilot study involving a single school district.

To determine whether children requiring special education represent a high-risk group for identifying Tourette's syndrome (TS), we performed direct examinations for the presence of tics in 35 special education and 35 regular classroom students from a single school district. Of the special education students, nine (26%) had definite or probable tics as compared with only two (6%) of the regular classroom students. About one-third of the students with tics currently meet diagnostic criteria for TS and probably more will do so in the future. About one-half of the subjects with tics have evidence of obsessive-compulsive behavior (OCB) or an attention-deficit hyperactivity disorder (ADHD). For three randomly selected students with definite tics, direct examinations of first-degree relatives revealed the presence of tics in all families. Subjects to the limitations of this pilot study, we conclude that TS and related tic disorders are commonly associated with the need for special education in this single school district. TS might also be an important contributor to school problems in the childhood population at large and may be a highly prevalent condition. In addition, we conclude that childhood tics are associated with OCB and ADHD, are genetically determined, and are part of the TS clinical spectrum.

Adolescent↗
PubMed1992

Stopping the circadian pacemaker with inhibitors of protein synthesis.

The requirement for protein synthesis in the mechanism of a circadian pacemaker was investigated by using inhibitors of protein synthesis. Continuous treatment of the ocular circadian pacemaker of the mollusc Bulla gouldiana with anisomycin or cycloheximide substantially lengthened (up to 39 and 52 hr, respectively) the free-running period of the rhythm. To determine whether high concentrations of inhibitor could stop the pacemaker, long pulse treatments of various durations (up to 44 hr) were applied and the subsequent phase of the rhythm was assayed. The observed phases of the rhythm after the treatments were a function of the time of the end of the treatment pulse, but only for treatments which spanned subjective dawn. The results provide evidence that protein synthesis is required in a phase-dependent manner for motion of the circadian pacemaker to continue.

Action Potentials↗
PubMed1990

The molecular genetic analysis of haemophilia A; characterization of six partial deletions in the factor VIII gene.

In a survey of 528 unrelated haemophilia A patients, six partial deletions of the factor VIII (FVIII) gene were detected by Southern blotting. These deletions were further mapped by a combination of Southern blotting and polymerase chain reaction amplification and found to vary in length between 4.7 kb and 57 kb. The frequency of detectable FVIII gene deletions (about 1%) frequency of detectable FVIII gene deletions (about 1%) is thus considerably lower than previously reported. Statistical analysis of currently available data did not provide any evidence for a deletion "'hotspot". Four of the six deletion patients reported here possessed inhibitors. Taken together with previous data, deletion of the FVIII gene was found to be associated with an approximately five-fold higher risk of developing inhibitors compared with other severe haemophiliacs without gene deletions.

Blotting, Southern↗
PubMed1972

Juvenile hormone induction of esterases: a mechanism for the regulation of juvenile hormone titer.

Within a few hours after injection of juvenile hormone into Hyalophora gloveri pupae, several fast-migrating carboxylesterases (EC 3.1.1.1) that are sensitive to diisopropylfluorophosphate appear in the hemolymph. Treatment of the pupae with puromycin or actinomycin D prevents the appearance of these hemolymph enzymes, suggesting de novo synthesis of the carboxylesterases. Of the several other compounds investigated, only a potent mimic of the juvenile hormone is able to induce these enzymes. When the induced enzymes are incubated in vitro with (14)C-labeled juvenile hormone, the hormone is rapidly and efficiently degraded. It is suggested that these induced carboxylesterases play an important role in the regulation of juvenile hormone titer.

Animals↗
PubMedDate not supplied

A clockwork organ.

The vertebrate circadian clock was thought to be highly localized to specific anatomical structures: the mammalian suprachiasmatic nucleus (SCN), and the retina and pineal gland in lower vertebrates. However, recent findings in the zebrafish, rat and in cultured cells have suggested that the vertebrate circadian timing system may in fact be highly distributed, with most if not all cells containing a clock. Our understanding of the clock mechanism has progressed extensively through the use of mutant screening and forward genetic approaches. The first vertebrate clock gene was identified only a few years ago in the mouse by such an approach. More recently, using a syntenic comparative genetic approach, the molecular basis of the the tau mutation in the hamster was determined. The tau gene in the hamster appears to encode casein kinase 1 epsilon, a protein previously shown to be important for PER protein turnover in the Drosophila circadian system. A number of additional clock genes have now been described. These proteins appear to play central roles in the transcription-translation negative feedback loop responsible for clock function. Post-translational modification, protein dimerization and nuclear transport all appear to be essential features of how clocks are thought to tick.

Animals↗