实验基础

An ideal biochemical source of a defined chromatin template assembled in vivo is the SV40 minichromosomes (see Note 1). At all stages in the viral lytic cycle, SV40 DNA is complexed with cellular histone and nonhistone proteins to form the episomal chromatin structure called a minichromosome ...

In vitro studies on nucleosome core particles (NCPs) and nucleosomes have generally been limited to the use of histone proteins isolated from chromatin. Numerous reliable and well-established methods have been described of obtaining single histone proteins in significant quant ...

In chromatin, each nucleosome contains two specific binding sites for nonhistone chromosomal proteins HMG-14 or HMG-17; however, because the amount of these proteins in the nucleus is limited, only a small fraction of the nucleosomes contain these proteins. A central question on the role of H ...

Eukaryotic genome replication implies that both the DNA and the associated proteins must be duplicated during each S phase. Model systems such as the simian virus 40 (SV40) in vitro replication system have provided important information on the mechanism of DNA replication in mammalian cel ...

In vivo DNA is associated with the proteins that constitute chromatin. This means that, any process that requires access to the genetic material must do so within the context of chromatin. It is now clear that there is a complex cellular machinery dedicated to regulating chromatin structure and t ...

Histones are the predominant protein component of chromatin and are subject to a variety of post-translational modifications. Of these, acetylation of the amino-terminal tails of core histones is most intensively studied and is linked to chromatin assembly, the regulation of gene exp ...

A central problem in the control of eukaryotic gene expression is how the compaction of DNA in chromatin is overcome to allow the initiation and elongation of transcription (1–6). Current studies reveal that multiple mechanisms are involved in counteracting chromatin-mediated repre ...

Treatment of nuclei with limited amounts of DNaseI can be used to reveal sites in chromatin that are hypersensitive (HS) to the nuclease (1,2). DNaseI HS sites are thought to correspond to sites where the regular nucleosome structure is perturbed, e.g., by binding of proteins to chromatin such that DNA ...

Posttranslational acetylation of the core histone amino-terminal tails correlates with both chromatin assembly and gene expression. This energy-intensive and reversible process is mediated by the opposing activities of histone acetyltransferase (HAT) and deacetylase ( ...

The structural and functional subunits of chromatin are nucleosome cores. In a nucleosome core 145 bp of DNA are coiled around the outer surface of an octamer of histone proteins which consists of a tetramer of 2(H3�H4) and two H2A�H2B dimers (1). DNA extending from the nucleosome core to the next nucleo ...

This chapter addresses the application of ribozymes in studying an important area of cancer biology, the process of metastasis. An example is given on how ribozymes can be used to study the role in this process of one particular gene. Initially, a brief introduction to the biology of cancer metastas ...

Inflammation and tissue injury are characterized by a massive infiltration of mononuclear cells responsible for the production of a variety of cytokines that alter the biosynthetic repertoire of the connective tissue cells (1,2). Recently, experiments in animal models of autoimmu ...

The use of oligodeoxynucleotides (ODN) to disrupt gene function has been studied in a variety of in vitro and in vivo systems (1–14). Antigene, antisense, ribozyme, and aptamer nucleic acid molecules have been shown in numerous model systems to effect DNA, RNA, or protein targets, or physiologic pr ...

It is often found that tumors have chromosomal translocation, especially in hematological malignancies. A specific chromosomal translocation is related to a specific type of disease. Recent studies have revealed that genetic aberrations originated from these chromosomal tr ...

The hammerhead ribozyme was discovered as a self-cleaving RNA molecule in certain plant viroids and satellite RNAs (1). Shortly after its conserved features were defined (2,3), the hammerhead was shown to be able to act as a true enzyme, cleaving multiple substrates in a bimolecular reaction (4). ...

The significance of blood vessel formation (angiogenesis) for the growth of solid tumors and ultimately their metastatic spread has gained wide acceptance. The close relationship between tumor angiogenesis and metastasis has been demonstrated in a series of correlative clinical ...

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder that results in dementia, and ultimately in the death of its victims. In the United States, AD is the fourth leading cause of death, and without a forseeable means of prevention, or effective treatment this will likely incre ...

The discovery of enzymatic RNA molecules, termed ribozymes (1,2), fundamentally changed our view of the function of RNA in chemistry, biology, and medicine. Naturally occurring enzymatic RNA molecules catalyze sequence-specific RNA processing. The sequence specificity is dete ...

The enzymatic activity of RNA molecules has been a source of surprises ever since 1981 when Cech discovered this new aspect of RNA function (1,2). Its role as messenger and structural component in the translational process was suddenly transcended, and it is now evident that various RNA molecules ...

The study of cellular function of a particular gene can be advanced by either specifically eliminating or blocking the function of its gene product. Although antisense RNA has frequently been used for this purpose, the hammerhead ribozyme has recently drawn a great deal of attention due to its po ...