Cell Cycle Control and Dysregulation Protocols focuses on emerging methodologies for studying the cell cycle, kinases, and kinase inhibitors. It addresses the issue of gene expression in vivo and in vitro, the analysis of cyclin-dependent kinase inhibitors, protein degradation mediated by the proteosome, the analysis of the transformed cell phenotype, and innovative techniques to detect apoptosis. Because there are already many manuals and protocols available, along with commercial kits and reagents, a variety of the more common techniques have not been included in our book. The protocols described, based on rather sophisticated techniques for in vivo and in vitro studies, consist of molecular biology, biochemistry, and various types of immunoassays. Indeed, the authors have successfully accomplished an arduous task by presenting several topics in the simplest possible manner. We are confident that Cell Cycle Control and Dysregulation Protocols will facilitate and optimize the work of practical scientists involved in researching the cell cycle. We greatly acknowledge the extraordinary contribution of the authors in writing this book.

The fundamental question of how cells grow and divide has perplexed biologists since the development of the cell theory in the mid-19th century, when it was recognized by Virchow and others that “all cells come from cells.” In recent years, considerable effort has been applied to the identification of the basic molecules and mechanisms that regulate the cell cycle in a number of different organisms. Such studies have led to the elucidation of the central paradigms that underpin eukaryotic cell cycle control, for which Lee Hartwell, Tim Hunt, and Paul Nurse were jointly awarded the Nobel Prize for Medicine and Physiology in 2001 in recognition of their seminal contributions to this field. The importance of understanding the fundamental mechanisms that modulate cell division has been reiterated by relatively recent discoveries of links between cell cycle control and DNA repair, growth, cellular metabolism, development, and cell death. This new phase of integrated cell cycle research provides further challenges and opportunities to the biological and medical worlds in applying these basic concepts to understanding the etiology of cancer and other proliferative diseases.

This book is a state-of-the-art summary of the latest achievements in cell cycle control research with an outlook on the effect of these findings on cancer research. The chapters are written by internationally leading experts in the field. They provide an updated view on how the cell cycle is regulated in vivo, and about the involvement of cell cycle regulators in cancer.

Intracellular checkpoint controls constitute a network of signal transd- tion pathways that protect cells from external stresses and internal errors. Ext- nal stresses can be generated by the continuous assault of DNA-damaging agents, such as environmental mutagens, ultraviolet (UV) light, ionizing radiation, or the reactive oxygen species that can arise during normal cellular metabolism. In response to any of these assaults on the integrity of the genome, the activation of the network of checkpoint control pathways can lead to diverse cellular responses, such as cell cycle arrest, DNA repair, or elimination of the cell by cell death (apoptosis) if the damage cannot be repaired. Moreover, internal errors can occur during the highly orchestrated replication of the cellular genome and its distribution into daughter cells. Here, the temporal order of these cell cycle events must be strictly enforced—for example, to ensure that DNA replication is c- plete and occurs only once before cell division, or to monitor mitotic spindle assembly, and to prevent exit from mitosis until chromosome segregation has been completed. Thus, well functioning checkpoint mechanisms are central to the maintenance of genomic integrity and the basic viability of cells and, the- fore, are essential for proper development and survival. The importance of proper functioning of checkpoints becomes plainly obvious under conditions in which this control network malfunctions and fails. Depending on the severity and timing, failure of this machinery can lead to embryonic lethality, genetic diseases, and cancer.

A collection of cutting-edge techniques for studying ubiquitin-dependent protein degradation via the proteasome. The topics covered range broadly from basic biochemistry to cellular assays to discovery techniques using mass spectrometric analysis. These biochemical and cellular methods are necessary to explore the ubiquitin-proteasome system and ubiquitin-proteasome-dependent functions. State-of-the-art and user-friendly, Ubiquitin-Proteasome Protocols offers novice and experienced bench scientists alike a thorough compendium of readily reproducible techniques that will accelerate discovery, enhance productivity, and permit manipulation of the system for varied research purposes.

A cutting-edge collection of basic and state-of-the-art methods optimized for investigating the molecular biology of this class of retrovirus. These readily reproducible techniques range from methods for the isolation and detection of human retroviruses to cutting-edge methods for exploring the interplay between the viruses and the host. Here, the researcher will find up-to-date techniques for the isolation and propagation of HIV, HTLV, and foamy virus from a variety of sources. There are also assays for determining the cell tropism of HIV-1, the coreceptor usage of HIV-1, and human gene expression with HIV-1 infection by microarrays, as well as for phenotyping HIV-1 infected monocytes and examining their fitness. Highlights include the detection and quantification of HIV-1 in resting CD4+, a new cloning system for making recombinent virus, cDNA microarrays, and the determination of genetic polymorphisms in two recently identified HIV-1 co-factors that are critical for HIV-1 infection.

A collection of new reviews and protocols from leading experts in cell cycle regulation, Cell Cycle Control: Mechanisms and Protocols, Second Edition presents a comprehensive guide to recent technical and theoretical advancements in the field. Beginning with the overviews of various cell cycle regulations, this title presents the most current protocols and state-of-the-art techniques used to generate latest findings in cell cycle regulation, such as protocols to analyze cell cycle events and molecules. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Cell Cycle Control: Mechanisms and Protocols, Second Edition will be a valuable resource for a wide audience, ranging from the experienced cell cycle researchers looking for new approaches to the junior graduate students giving their first steps in cell cycle research.