细胞技术

An important component of the stress response, as well as many other biological systems, is the change in relative concentration of specific mRNAs following induction (1). Conventional techniques of differential screening may be sufficient for the detection and identification of so ...

The characteristics of an organism or tissue are determined by the genes expressed within it. The determination of the genomic sequence of higher organisms, including humans, is now a real and attainable goal, as seen by the progress of the Human Genome Mapping Project. This information is needed ...

The heme oxygenase enzymes (HO-1 and HO-2) oxidize heme to biliver-din-IXα (BVIXα), releasing carbon monoxide (CO) and iron (Fig. 1). HO enzymes control the rate of heme degradation and, consequently, also control the redistribution of the heme iron (1). The CO generated from the HO reaction affects ...

Molecular chaperones function in a range of protein homeostatic events, including cotranslational protein folding, assembly and disassembly of protein complexes, and protein transport across membranes. Many molecular chaperones are also known as heat-shock proteins, which ...

Small Hsp’s (sHsp’s) are an ubiquitous but diverse class of proteins that differ from other Hsp families in that only certain short-sequence motifs, the so-called α-crystallin domains and some sequence in the N-terminal parts, are conserved. Characteristic features in common are their low m ...

The p53 protein (reviewed in refs. 1–5) is a latent transcription factor that is activated in response to a variety of cellular stresses, including DNA damage, mitotic spindle damage, heat shock, hypoxia, cytokines, metabolic changes, viral infection, and activated oncogenes, and is consid ...

Small heat-shock proteins (sHsps) are involved in diverse biological phenomena and can act as molecular chaperones in vitro (reviewed in ref. 1, see also Chapter 25). At the posttranslational level, several modifications of sHsp’ have been detected, including phosphorylation, deamid ...

Multicellular animals are exposed routinely to oxidizing chemicals and radiation from the environment, as well as endogenous metabolic by-products that can damage DNA and proteins over the life-span of the cell. Such damage may contribute to tissue injury, promote aging, and is implicat ...

The first preliminary characterization of any newly isolated DNA fragment usually involves restriction site mapping. For this, a family of bacterial enzymes called restriction endonucleases, or restriction enzymes, are utilized. These enzymes recognize specific sequences ...

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 ...