细胞功能测定

The 20S proteasome is part of a larger complex, the 26S proteasome, that is implicated in the ATP-dependent degradation of multiubiquitin-conjugated proteins (1). About 80% of intracellular protein breakdown occurs via the ubiquitinproteasome system (UPS). Key proteins such as tran ...

An emerging theme in neurodegenerative diseases is the aggregation of proteins as inclusions in neural cells. Their presence is a useful tool in the differential diagnosis of the particular illness, although in no instance is the specific role of the inclusions in disease pathogenesis und ...

A method for isolation of constitutive heterochromatin (chromocenters) from nuclei of mouse liver cells is described. This method is based on the higher resistance of chromocenters to low ionic strength treatment as compared with that of nucleoli and euchromatin. The method allows sep ...

The perinucleolar compartment (PNC) is a dynamic, irregularly shaped, and electron-dense nuclear structure that is physically associated with the nucleolus (1). It is found predominantly in transformed cells and various cancer tissues, and rarely in normal cells (1). The components of ...

The dynamics of proteins play a key role in the organization and control of nuclear functions. Techniques were developed recently to observe the movement and interactions of proteins in living cells; time-lapse microscopy using fluorescent-tagged proteins gives access to observat ...

Here we describe a new, rapid method for isolating nucleoli from Ehrlich tumor cells that preserves their morphological integrity and high transcriptional activity. Until now, methods for isolation of nucleoli were generally assumed to empty one of their three main compartments, the fi ...

The nucleus is a complex volume constituted of numerous subcompartments in which specific functions take place due to a specific spatial organization of their molecular components. To understand how these molecules are spatially organized within these machineries, it is necessary ...

Dinoflagellates are unicellular eukaryotic organisms whose nuclear structure, chromosome architecture, chromatin organization, DNA composition, and mitosis show original features. It has been necessary to adapt techniques and to create innovative methods for growing c ...

The vast evolutionary distance between the Opisthokonta (animals and yeast) and the excavata (a major group of protists, including Giardia and Trypanosoma) presents a significant challenge to in silico functional genomics and ortholog identification. Subcellular proteomic i ...

We describe a method for isolating nuclei from cultured Arabidopsis cells. The same method can be used to further isolate nucleoli. Cell walls are first digested to yield protoplasts, which are purified by flotation on a Percoll gradient. Mechanical homogenisation is used to release nuclei, ...

The giant nucleus or germinal vesicle (GV) of amphibian oocytes presents a remarkable opportunity to examine nuclear structures in unprecedented levels of detail. By making use of spread preparations of GVs, it is possible to investigate the structure and function of transcription uni ...

Probing chromatin structure with nucleases is a well-established method for determining the accessibility of DNA to gene regulatory proteins and measuring competency for transcription. A hallmark of many silent genes is the presence of translationally positioned nucleosomes ...

The nuclear envelope is a complex membrane-protein system that is notoriously difficult to purify because it has many connections to both nuclear and cytoplasmic components. This difficulty is compounded by the fact that the nature of these connections vary in different cell types, and so m ...

Many of the chapters in this volume are concerned with processes or structures inside the nucleus, and it is relevant to consider the properties of their environment, or rather of the multiple different and specific environments that must exist in local regions of the highly heterogeneous int ...

DNA double-strand breaks (DSBs) are among the most dangerous types of DNA damage. Unrepaired, DSBs may lead to cell death, and when misrejoined, they can result in potentially carcinogenic chromosome rearrangements. The induction of DSBs and their repair take place in a chromatin microenv ...

Epigenetic mechanisms lead to the stable regulation of gene expression without alteration of DNA and trigger initiation and/or maintenance of cell-type-specific transcriptional profiles. Indeed, modulation of chromatin structure and the global 3D organization of the genome ...

A myriad of cellular processes instigated by growth factors are mediated by cell surface-associated receptor tyrosine kinases (RTKs). Subsequent downstream activation of signaling cascades, as well as their crosstalk, endows specificity in terms of the phenotypic outcome, e.g., c ...

A complex intracellular signaling network mediates the multiple biological activities of G-protein-coupled receptors (GPCRs). Among them, monomeric GTPases and a family of closely related proline-targeted serine–threonine kinases, collectively known as Mitogen-Act ...

Mitogen-activated protein kinase (MAPK) signaling influences a variety of cellular responses, ranging from stimulation of cell proliferation to induction of senescence and/or apoptosis. Ca2+ is a ubiquitous intracellular signaling molecule that controls multiple proces ...

ERK-MAPK is activated by dual phosphorylation of its activation loop TEY motif by the MEK-MAPKK. ERK cytoplasmic activity should be measured by assaying both the level of dually phosphorylated ERK and the level of phosphorylated substrate. We describe two complementary methods for quan ...