细胞功能测定

Mitochondrial DNA (mtDNA) in animal cells is organized into clusters of 5–7 genomes referred to as nucleoids. Contrary to the notion that mtDNA is largely free of bound proteins, these structures are nearly as rich in protein as nuclear chromatin. While the purification of intact, membrane-bo ...

The bulk of ATP consumed by various cellular processes in higher eukaryotes is normally produced by five multimeric protein complexes (I–V) embedded within the inner mitochondrial membrane, in a process known as oxidative phosphorylation (OXPHOS). Maintenance of energy homeosta ...

In higher vertebrates, the DNA of mitochondria takes the form of circular molecules of approximately 16 kbp. These circles are arranged in multigenomic nucleoprotein complexes or nucleoids. It is envisaged that nucleoid superstructure makes a critical contribution to the twin proc ...

The mitochondrion is the eukaryotic organelle that carries out oxidative phosphorylation, fulfilling cellular requirements for ATP production. Disruption of mitochondrial energy metabolism can occur by genetic and biochemical mechanisms involving nuclear-encoded ...

DNA polymerase γ (pol γ) is the only DNA polymerase within the mitochondrion and is thus essential for replication and repair of mtDNA. POLG, the gene encoding the catalytic subunit of pol γ, is a major locus for a wide spectrum of mitochondrial diseases with more than 100 known disease mutations. Thus, we n ...

In this chapter, we present a streamlined purification for the production of near-homogeneous and high yield recombinant forms of the human mitochondrial DNA helicase. Minimizing the number of steps and the time elapsed for purification of this enzyme facilitates studies of its struct ...

Mitochondrial DNA (mtDNA) maintenance can be and has been studied in a wide variety of organisms, some more tractable than others. We use human and mouse cell culture models to study proteins and mechanisms of mtDNA replication. Recent advances in cell culture systems allow us to streamline the an ...

Characterization of the basic transcription machinery of mammalian mitochondrial DNA has been greatly supported by the availability of pure recombinant mitochondrial RNA polymerase (mtRNAP) and accessory factors, which allowed to develop a reconstituted in vitro transcri ...

Kinetochores are essential for the proper positioning, movement and segregation of chromosomes on spindle microtubules. Live cell analyses of kinetochore movements on the spindle provide an important tool for dissecting the molecular machinery underlying kinetochore-ba ...

In neurons, the molecular machinery for axonal growth and navigation is localized to the growth cone region, whereas tubulin is synthesized primarily in the cell body. Because diffusion serves as an efficient transport mechanism only for very short distances, tubulin has to be actively tra ...

Host microtubules and motor proteins are crucial to the intracellular transport of a number of viruses. Disruption of microtubules or suppression of motor functions can remarkably inhibit the movement of viruses in host cells. It is now known that incoming viruses use motor proteins to trav ...

Microtubule-actin interactions are fundamental to many cellular processes such as cytokinesis and cellular locomotion. Investigating the mechanism of microtubule-actin interactions is the key to understand the cellular morphogenesis and related pathological proce ...

The mitotic spindle is an important target for cancer chemotherapy. The main protein target for drugs in clinical use is tubulin, the building block of microtubules. In recent years, other proteins of the mitotic spindle have been identified as potential targets for the development of more spe ...

The microtubule-targeting agents have made significant contributions to cancer therapy over the past 50 years. The vinca alkaloids and taxanes have been used to treat a broad range of malignancies, including leukemias and lymphomas and many types of solid tumors. The taxanes have been freq ...

A method is described for measurement of the circadian activity rhythms of up to 150 larval zebrafish simultaneously with a single video image analysis system. Most of the required equipment and software are commercially available, although some components are custom-built.

Many of the behavioral parameters exhibited by an organism show daily fluctuations. These may persist under constant environmental conditions, demonstrating that they are governed by an endogenous (circadian) clock. The monitoring of locomotor activity in rodents is probably one ...

The unicellular cyanobacterium Synechococcus elongatus PCC 7942 is the model organism for studying prokaryotic circadian rhythms. Although S. elongatus does not display an easily measurable overt circadian behavior, its gene expression is under circadian control; hence, a “be ...

Arabidopsis thaliana is the model organism for the study of the higher plant circadian clock. The physiological change in position of young leaves and cotyledons in Arabidopsis seedlings reveals an overt circadian rhythm. Measuring these leaf movements provides a simple and reliable a ...

Synechococcus elongatus PCC 7942 is an excellent model system for studying the molecular mechanism of the circadian clock in cyanobacteria. The “plastic” genetic characteristics of this organism have facilitated the development of various methods for mutagenesis of its chromos ...

The study of circadian clock function in Drosophila relies heavily on the analysis of rhythmic gene expression. Typically, individuals or groups of flies collected during a specific time of the circadian day need to be sacrificed, followed by the extraction of clock gene products. This proce ...