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Biomedical subjects

W F Marshall

Publications and source records attributed to W F Marshall.

At least 19 recordsLinked to original sources
PubMed2001

Intraflagellar transport balances continuous turnover of outer doublet microtubules: implications for flagellar length control.

A central question in cell biology is how cells determine the size of their organelles. Flagellar length control is a convenient system for studying organelle size regulation. Mechanistic models proposed for flagellar length regulation have been constrained by the assumption that flagella are static structures once they are assembled. However, recent work has shown that flagella are dynamic and are constantly turning over. We have determined that this turnover occurs at the flagellar tips, and that the assembly portion of the turnover is mediated by intraflagellar transport (IFT). Blocking IFT inhibits the incorporation of tubulin at the flagellar tips and causes the flagella to resorb. These results lead to a simple steady-state model for flagellar length regulation by which a balance of assembly and disassembly can effectively regulate flagellar length.

Animals↗
PubMed2001

Centrioles take center stage.

Centrioles are among the most beautiful and mysterious of all cell organelles. Although the ultrastructure of centrioles has been studied in great detail ever since the advent of electron microscopy, these studies raised as many questions as they answered, and for a long time both the function and mode of duplication of centrioles remained controversial. It is now clear that centrioles play an important role in cell division, although cells have backup mechanisms for dividing if centrioles are missing. The recent identification of proteins comprising the different ultrastructural features of centrioles has proven that these are not just figments of the imagination but distinct components of a large and complex protein machine. Finally, genetic and biochemical studies have begun to identify the signals that regulate centriole duplication and coordinate the centriole cycle with the cell cycle.

Animals↗
PubMed2001

Chromosome elasticity and mitotic polar ejection force measured in living Drosophila embryos by four-dimensional microscopy-based motion analysis.

BACKGROUND: Mitosis involves the interaction of many different components, including chromatin, microtubules, and motor proteins. Dissecting the mechanics of mitosis requires methods of studying not just each component in isolation, but also the entire ensemble of components in its full complexity in genetically tractable model organisms. RESULTS: We have developed a mathematical framework for analyzing motion in four-dimensional microscopy data sets that allows us to measure elasticity, viscosity, and forces by tracking the conformational movements of mitotic chromosomes. We have used this approach to measure, for the first time, the basic biophysical parameters of mitosis in wild-type Drosophila melanogaster embryos. We found that Drosophila embryo chromosomes are significantly less rigid than the much larger chromosomes of vertebrates. Anaphase kinetochore force and nucleoplasmic viscosity were comparable with previous estimates in other species. Motion analysis also allowed us to measure the magnitude of the polar ejection force exerted on chromosome arms during metaphase by individual microtubules. We find the magnitude of this force to be approximately 1 pN, a number consistent with force generation either by collision of growing microtubules with chromosomes or by single kinesin motors. CONCLUSIONS: Motion analysis allows noninvasive mechanical measurements to be made in complex systems. This approach should allow the functional effects of Drosophila mitotic mutants on chromosome condensation, kinetochore forces, and the polar ejection force to be determined.

Algorithms↗
PubMed2001

Kinetics and regulation of de novo centriole assembly. Implications for the mechanism of centriole duplication.

BACKGROUND: Centriole duplication is a key step in the cell cycle whose mechanism is completely unknown. Why new centrioles always form next to preexisting ones is a fundamental question. The simplest model is that preexisting centrioles nucleate the assembly of new centrioles, and that although centrioles can in some cases form de novo without this nucleation, the de novo assembly mechanism should be too slow to compete with normal duplication in order to maintain fidelity of centriole duplication. RESULTS: We have measured the rate of de novo centriole assembly in vegetatively dividing cells that normally always contain centrioles. By using mutants of Chlamydomonas that are defective in centriole segregation, we obtained viable centrioleless cells that continue to divide, and find that within a single generation, 50% of these cells reacquire new centrioles by de novo assembly. This suggests that the rate of de novo assembly is approximately half the rate of templated duplication. A mutation in the VFL3 gene causes a complete loss of the templated assembly pathway without eliminating de novo assembly. A mutation in the centrin gene also reduced the rate of templated assembly. CONCLUSIONS: These results suggest that there are two pathways for centriole assembly, namely a templated pathway that requires preexisting centrioles to nucleate new centriole assembly, and a de novo assembly pathway that is normally turned off when centrioles are present.

Animals↗
PubMed2001

Bad neighbors.

Explore the source record for details and available documents.

Chromosome Inversion↗
PubMed2000

How centrioles work: lessons from green yeast.

Centrioles are the organizing centers around which centrosomes assemble. Despite a century of study, the molecular details of centriole function and assembly remain largely unknown. Recent work has exploited the unique advantages of unicellular algae to reveal proteins that play central roles in centriole biology.

Animals↗
PubMed1999

Cell division: The renaissance of the centriole.

Centrioles are located at the center of the cytoskeleton and duplicate exactly once per cell cycle. Recent studies suggest that centrioles are required for the organization of a functional centrosome and that centriole assembly requires both gamma- and delta-tubulin.

Animals↗
PubMed1999

The cephalosporins.

The cephalosporins are a large group of related beta-lactam antimicrobial agents. Favorable attributes of the cephalosporins include low rates of toxicity, relatively broad spectrum of activity, and ease of administration. Various cephalosporins are effective for treatment of many conditions, including pneumonia, skin and soft tissue infections, bacteremia, and meningitis. Differences among the numerous cephalosporin antimicrobial agents are sometimes subtle; however, an understanding of these differences is essential for optimal use of these agents. As a result of widespread use of cephalosporins, bacterial resistance to these drugs is increasingly common. New, fourth-generation agents (such as cefepime) offer an alternative for the treatment of infections caused by some drug-resistant microorganisms.

Administration, Oral↗
PubMed1998

Homologous chromosome pairing in Drosophila melanogaster proceeds through multiple independent initiations.

The dynamics by which homologous chromosomes pair is currently unknown. Here, we use fluorescence in situ hybridization in combination with three-dimensional optical microscopy to show that homologous pairing of the somatic chromosome arm 2L in Drosophila occurs by independent initiation of pairing at discrete loci rather than by a processive zippering of sites along the length of chromosome. By evaluating the pairing frequencies of 11 loci on chromosome arm 2L over several timepoints during Drosophila embryonic development, we show that all 11 loci are paired very early in Drosophila development, within 13 h after egg deposition. To elucidate whether such pairing occurs by directed or undirected motion, we analyzed the pairing kinetics of histone loci during nuclear cycle 14. By measuring changes of nuclear length and correlating these changes with progression of time during cycle 14, we were able to express the pairing frequency and distance between homologous loci as a function of time. Comparing the experimentally determined dynamics of pairing to simulations based on previously proposed models of pairing motion, we show that the observed pairing kinetics are most consistent with a constrained random walk model and not consistent with a directed motion model. Thus, we conclude that simple random contacts through diffusion could suffice to allow pairing of homologous sites.

Animals↗
PubMed1997

Interphase chromosomes undergo constrained diffusional motion in living cells.

BACKGROUND: Structural studies of fixed cells have revealed that interphase chromosomes are highly organized into specific arrangements in the nucleus, and have led to a picture of the nucleus as a static structure with immobile chromosomes held in fixed positions, an impression apparently confirmed by recent photobleaching studies. Functional studies of chromosome behavior, however, suggest that many essential processes, such as recombination, require interphase chromosomes to move around within the nucleus. RESULTS: To reconcile these contradictory views, we exploited methods for tagging specific chromosome sites in living cells of Saccharomyces cerevisiae with green fluorescent protein and in Drosophila melanogaster with fluorescently labeled topoisomerase ll. Combining these techniques with submicrometer single-particle tracking, we directly measured the motion of interphase chromatin, at high resolution and in three dimensions. We found that chromatin does indeed undergo significant diffusive motion within the nucleus, but this motion is constrained such that a given chromatin segment is free to move within only a limited subregion of the nucleus. Chromatin diffusion was found to be insensitive to metabolic inhibitors, suggesting that it results from classical Brownian motion rather than from active motility. Nocodazole greatly reduced chromatin confinement, suggesting a role for the cytoskeleton in the maintenance of nuclear architecture. CONCLUSIONS: We conclude that chromatin is free to undergo substantial Brownian motion, but that a given chromatin segment is confined to a subregion of the nucleus. This constrained diffusion is consistent with a highly defined nuclear architecture, but also allows enough motion for processes requiring chromosome motility to take place. These results lead to a model for the regulation of chromosome interactions by nuclear architecture.

Animals↗
PubMed1997

Mitosis in living budding yeast: anaphase A but no metaphase plate.

Chromosome movements and spindle dynamics were visualized in living cells of the budding yeast Saccharomyces cerevisiae. Individual chromosomal loci were detected by expression of a protein fusion between green fluorescent protein (GFP) and the Lac repressor, which bound to an array of Lac operator binding sites integrated into the chromosome. Spindle microtubules were detected by expression of a protein fusion between GFP and Tub1, the major alpha tubulin. Spindle elongation and chromosome separation exhibited biphasic kinetics, and centromeres separated before telomeres. Budding yeast did not exhibit a conventional metaphase chromosome alignment but did show anaphase A, movement of the chromosomes to the poles.

Anaphase↗
PubMed1997

Prophylaxis of cytomegalovirus infection in liver transplantation: a randomized trial comparing a combination of ganciclovir and acyclovir to acyclovir. NIDDK Liver Transplantation Database.

BACKGROUND: The optimal prophylactic regimen to prevent cytomegalovirus (CMV) infection and disease in orthotopic liver-transplant patients remains to be established. We tested whether a combination of intravenous ganciclovir (GCV) followed by high dosages of oral acyclovir (ACV) for 4 months provided a higher degree of protection from CMV than oral ACV alone. METHODS: One hundred sixty-seven liver-transplant recipients were randomized to receive 120 days of antiviral treatment starting at the time of transplantation consisting of either ACV 800 mg orally four times daily (n=84) or 14 days of GCV 5 mg/kg intravenously every 12 hr followed by oral ACV 800 mg four times daily (n=83). Prospective laboratory and clinical surveillance was performed to determine primary endpoints (onset of CMV infection and CMV disease) and secondary endpoints (rates of fungal and bacterial infection, allograft rejection, and survival after transplantation). One-year event rates are presented as cumulative percentages. RESULTS: During the first year after transplantation, CMV infection developed in 57% of patients treated with ACV and in 37% of patients treated with GCV + ACV (P=0.001). CMV disease developed in 23% of patients treated with ACV and in 11% of patients treated with GCV + ACV (P=0.03). In seronegative recipients of allografts from CMV-seropositive donors (D+/R-), CMV disease developed in 58% of patients treated with ACV and in 25% of patients treated with GCV + ACV (P=0.04). In the D+/R- group, 54% of patients treated with ACV and 17% of patients treated with GCV + ACV developed infection with Candida albicans (P=0.05). CONCLUSIONS: Prophylaxis of CMV infection in liver-transplant patients with 14 days of intravenous GCV followed by high-dosage oral ACV is more effective than high-dosage oral ACV alone at reducing CMV infection and disease, even for patients in the D+/R- CMV serological group.

Acyclovir↗
PubMed1997

Telomeres cluster de novo before the initiation of synapsis: a three-dimensional spatial analysis of telomere positions before and during meiotic prophase.

We have analyzed the progressive changes in the spatial distribution of telomeres during meiosis using three-dimensional, high resolution fluorescence microscopy. Fixed meiotic cells of maize (Zea mays L.) were subjected to in situ hybridization under conditions that preserved chromosome structure, allowing identification of stage-dependent changes in telomere arrangements. We found that nuclei at the last somatic prophase before meiosis exhibit a nonrandom, polarized chromosome organization resulting in a loose grouping of telomeres. Quantitative measurements on the spatial arrangements of telomeres revealed that, as cells passed through premeiotic interphase and into leptotene, there was an increase in the frequency of large telomere-to-telomere distances and a decrease in the bias toward peripheral localization of telomeres. By leptotene, there was no obvious evidence of telomere grouping, and the large, singular nucleolus was internally located, nearly concentric with the nucleus. At the end of leptotene, telomeres clustered de novo at the nuclear periphery, coincident with a displacement of the nucleolus to one side. The telomere cluster persisted throughout zygotene and into early pachytene. The nucleolus was adjacent to the cluster at zygotene. At the pachytene stage, telomeres rearranged again by dispersing throughout the nuclear periphery. The stage-dependent changes in telomere arrangements are suggestive of specific, active telomere-associated motility processes with meiotic functions. Thus, the formation of the cluster itself is an early event in the nuclear reorganizations associated with meiosis and may reflect a control point in the initiation of synapsis or crossing over.

Cell Nucleus↗