Understanding the details of how genetic information is expressed from the separate mitochondrial genome requires a detailed description of the properties of the mitochondrial RNA polymerase. This nuclear-encoded enzyme is necessary and sufficient for the transcription of all ...
Ribonucleic acid (RNA) interference triggered by double-stranded RNA has become a powerful tool for generating loss-of-function phenotypes. It is used to inactivate genes of interest and represents an elegant approach to genome functional analysis by reverse genetics. In Drosoph ...
Replication intermediates can be separated on agarose gels in two dimensions to reveal a wealth of data on mechanisms of DNA replication. When applied to mitochondrial DNA of higher vertebrates, this technique unearthed a host of unexpected findings, the full implications of which are sti ...
Ribonucleic acid (RNA) import into mitochondria occurs in a variety of organisms. In mammalian cells, several small RNAs are imported in a natural manner; transfer RNAs (tRNAs) can be imported in an artificial way, following expression of corresponding genes from another organism (yeast) ...
Mitochondrial biogenesis is an intricate process that requires the coordinated function of two separate genetic systems: one in the organelle and one in the nucleus. The study of mitochondria requires the analysis of both genetic systems and their protein products. We describe the gener ...
Amino acids are not only substrates for various metabolic pathways, but can also serve as signaling molecules controlling signal transduction pathways. One of these signaling pathways is mTOR-dependent and is activated by amino acids (leucine in particular) in synergy with insulin. A ...
Autophagy is a cellular homeostasis pathway used to sustain cellular anabolic needs during times of nutrient or energy deprivation. Autophagosomes sequester cytoplasmic constituents, including macromolecules such as long-lived proteins. Upon fusion of autophagosomes w ...
Insects such as the fruit fly Drosophila melanogaster, which fundamentally reorganize their body plan during metamorphosis, make extensive use of autophagy for their normal development and physiology. In the fruit fly, the hepatic/adipose organ known as the fat body accumulates nut ...
Recent studies of the molecular mechanism of autophagy have made available several marker proteins for autophagosomes. These marker proteins allow us to identify autophagic structures easily and accurately by fluorescent microscopy. The most widely used marker for autophagos ...
Autophagy is a physiological process functionally linked to cellular dynamics during starvation, cardiomyopathies, neurodegeneration, cellular immunity, and certain cancers. Although nearly 30 autophagy-related (ATG) genes have been identified and characterized, t ...
In this chapter, we explain different strategies to analyze the extracellular Hedgehog (Hh) morphogen distribution and Hh intracellular trafficking by immunohistochemistry techniques. For this purpose, it has been very useful to have a transgenic fly line that expresses a Hh-green ...
Hedgehog (Hh) family members are secreted proteins that can act at short and long range to direct cell fate decisions during developmental processes. In both Drosophila and vertebrates, the morphogenetic gradient of Hh must be tightly regulated for correct patterning. The posttransla ...
The identification of protein domains required for function is an important means of defining biochemical roles for a polypeptide. Our studies on regulatory proteins that function during the transition between mitosis and meiosis have extensively relied on targeted in vitro mutag ...
Genetic screens have been extraordinarily useful for the identification of protein components that function in a variety of cellular processes. For example, a number of proteins that are necessary for responding to deoxyribonucleic acid (DNA) damage were found in screens for loss-of- ...
The technique described in this chapter—gene targeting in cultured human cancer cells—brings a powerful tool to scientists studying the function of cell cycle control genes (1). This technology allows scientists to knock out genes in cultured human cells in an analogous fashion to the crea ...
The generation and analysis of mutants has had an essential role in the identification of genes involved in cell cycle control (1–3). In this regard, the genetic analysis of mutations has helped reveal the normal function of wild-type gene products as well as provided powerful insights into the in ...
Cell cycle phase-specific regulation of transcription is a major mechanism for the regulation of progress through the eukaryotic cell cycle. Hundreds of genes are known to be cell cycle regulated, including histones, cyclins, transcription factors, and genes for such cycle-specific ...
Fission yeast is a popular model organism for the study of the cell cycle. It grows quickly compared with other eukaryotic species; under normal conditions, a wild-type cell takes about 2.5–3 h to complete a cell cycle. A wild-type S. pombe cell has a rod shape, grows by elongation, and divides by medial fissi ...
The budding yeast Saccharomyces cerevisiae represents the eukaryotic model system in which the checkpoint concepts were initially developed (1). Whereas many arguments can be made in favor of the continued use of S. cerevisiae as a model organism for checkpoint studies, the ease of disting ...
Saccharomyces cerevisiae, the budding yeast, is widely used as a model eukaryote to study a large number of cellular processes including cell cycle regulation (1–4). Extensive genetic research in the last two decades has revealed that the basic mechanism of cell cycle control is highly conse ...
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