Advances in Molecular and Cell Biology – serie

Caveolae (latin for little caves) are small structures found at the surface of cells. They are responsible for the regulation of important metabolic pathway. As a consequence, they may play a critical role in several human diseases such as atherosclerosis, cancer, diabetes, and muscular dystrophies. This book analyzes the role and function of caveolae in these aspects and serves as the first textbook currently available on caveolae/caveolin.

This readable, comprehensive text covers endothelial biology from the fundamentals of structure and lung fluid balance physiology to state-of-the-art descriptions of the molecular mechanisms involved in the development of lung failure. The material and illustrations, provided by outstanding experts in their individual areas of research and clinical concentration, is artfully woven together to provide the reader with an integrated, in-depth, and up-to-date knowledge of endothelial function, vascular integrity, pulmonary function, and pathophysiology in respiratory failure.

This volume provides an overview of the cytoskeleton particularly on the fundamental role the cytoskeleton plays in the regulation of cell structure and function. This book represents new trends in cytoskeletal research that go beyond the traditional approach of identifying new proteins in the cytoskeleton, but actually define how these proteins interact with signaling pathways. While the major emphasis in this volume remains on the microfilament structure, some discussion has been included in this volume to illustrate the similarities and differences between the three cytoskeletal elements namely the actin microfilament, the intermediate filaments and the microtubules.

All living cells are surrounded by a lipidic membrane that isolates them from the often harsh environment. However, to take up nutrients, to excrete waste, and to communicate among each other, Nature has invented an incredibly diverse set of transmembrane transport proteins. Specialized transporters exist to shuttle electrically charged ions, positive cations like sodium or negative anions like chloride, across the membrane. In the recent years, tremendous progress has been made in the field of chloride transport. The present book presents the state of the art of this rapidly expanding and interest-gaining field of membrane transport. It is addressed at a broad medically, physiologically, biologically, and biophysically interested readership. Describes the state-of-the-art in anion transport research Written by leaders in the field Presents a timely discussion of this rapidly emerging and expanding field

Research on the cytochrome P450 family of genes has traditionally been dominated by forms participating in drug metabolism. This has occurred in spite of early discovery of steroid hydroxy lase P450 cytochromes in the adrenal gland. More recently, contributions on the characterization and regulation of P450 cytochromes involved in biosynthetic reactions have been found at the international meetings on cytochrome P450 and in the several books on the field. Key recognition that P450 cytochromes should be recognized in a physiological context was provided by an international meeting in Jerusalem in 1991 and the subsequent publication of the proceedings in the Journal of Steroid Biochemistry and Molecular Biology (43, number 8, 92). Like this meeting, this book seeks to place equal weight on the physiological processes that are controlled by the products of reactions at usually very selective cytochrome P450 forms. Each of the authors was asked to discuss the molecular regulation of these P450 forms. Each of the authors was asked to discuss the molecular regulation of these P450 forms in the light of these physiological processes. In some cases the physiological role of the cytochrome P450 and even the natural substrate are unresolved, but a pattern of strong endocrine regulation is indicative of a hidden function. As more and more low abundance P450 genes are uncovered, the need to address potential physiological activities becomes more pressing. It is almost infinitely more difficult to identify a physiological substrate than to clone a new form.

One prerequisite for the evolution of multicellular organisms was the invention of mechanisms by which cells could adhere to one another. At some point in our history, dividing cells no longer went their separate protozoic ways in the primordial oceans, but instead found that by maintaining an association, by sticking together but not fusing, numerous evolutionary advantages became possible. The subsequent development of specialized tissues and organs depended on the elaboration of incredibly sophisticated, regulatable cell-to-cell adhesion mechanisms which are known to operate in biological processes as diverse as the growth of the embryo, the immune response, the establishment of connections between nerve cells, and arteriosclerosis, to name just a few. Although we can only guess at the ancestral mechanisms that fostered the first primitive intercellular unions, some one billion years ago, we now recognize contemporary molecular "themes" with presumably ancient origins that mediate cell-cell interactions.

The chapters in this book serve as useful, thought-provoking, but not exhaustive, commentaries on contemporary topics within the broad field of cell adhesion. If the reader detects a slight tilt toward those adhesion molecules that function in the nervous system, this is merely a reflection of this editor's interests, biases, and of course, limitations.

A large number of newly-synthesized polypeptides must cross one or several intracellular membranes to reach their functional locations in the eukaryotic cell. The mechanisms of protein trafficking, in particular the post-translational targeting and membrane translocation of proteins, are of fundamental biological importance and are the focus of intensive research world-wide. For more than 15 years, mitochondria have served as the paradigm organelle system to study these processes. Although key questions, such as how precisely proteins cross a membrane, still remain to be answered, exciting progress has been made in understanding the basic pathways of protein import into mitochondria and the components involved. In addition to a fascinating richness and complexity in detail, the analysis of mitochondrial protein import has revealed mechanistic principles of general significance: Major discoveries include the demonstration of the requirement of an unfolded state for translocation and of the essential role of molecular chaperones on both sides of the membranes in maintaining a translocation-competent conformation and in protein folding after import. It is becoming clear how a polypeptide chain is "reeled" across the membrane in an ATP-dependent process by the functional cooperation of membrane proteins, presumably constituting part of a transmembrane channel, with peripheral components at the trans-side of the membrane.In this volume, eminent experts in the field take the time to review the central aspects of mitochondrial biogenesis. The logical order of the 16 chapters is determined by the sequence of steps during protein import, starting with the events taking place in the cytosol, followed by the recognition of targeting signals, the translocation of precursor proteins across the outer and inner membranes, their proteolytic processing and intramitochondrial sorting, and finally their folding and oligomeric assembly. In addition, the mechanisms involved in the export of mitochondrially encoded proteins as well as recent advances in understanding the division and inheritance of mitochondria will be discussed.

The revolution in biological research initiated by the demonstration that particular DNA molecules could be isolated, recombined in novel ways, and conveniently replicated to high copy number in vivo for further study, that is, the recombinant DNA era, has spawned many additional advances, both methodological and intellectual, that have enhanced our understanding of cellular processes to an astonishing degree. As part of the subsequent outpouring of information, research exploring the mechanisms of gene regulation, both in prokaryotes and eukaryotes (but particularly the latter), has been particularly well represented. Although no one technical approach can be said to have brought the filed to its current level of sophistication, the ability to map the interactions of trans-acting factors with their DNA recognition sequences to a high level of precision has certainly been one of the more important advances. This "footprinting" approach has become almost ubiquitous in gene regulatory studies; however, it is in its "in vivo" application that ambiguities, confusions, and inconsistencies that may arise from a purely "in vitro"-based approach can often be resolved and placed in their proper perspective. Put more simply, that an interaction can be demonstrated to occur between purified factors and a particular piece of DNA in a test tube does not, of course, say anything regarding whether such interactions are occurring in vivo. The ability to probe for such interactions as they occur inside cells, with due attention paid to the relevant developmental stage, or to the tissue specificity of the interaction being probed, has made in vivo footprinting approach an invaluable adjunct to the "gene jockey's" arsenal of weapons.

Recent advances in protein structural biology, coupled with new developments in human genetics, have opened the door to understanding the molecular basis of many metabolic, physiological, and developmental processes in human biology. Medical pathologies, and their chemical therapies, are increasingly being described at the molecular level. For single-gene diseases, and some multi-gene conditions, identification of highly correlated genes immediately leads to identification of covalent structures of the actual chemical agents of the disease, namely the protein gene products. Once the primary sequence of a protein is ascertained, structural biologists work to determine its three-dimensional, biologically active structure, or to predict its probable fold and/or function by comparison to the data base of known protein structures. Similarly, three-dimensional structures of proteins produced by microbiological pathogens are the subject of intense study, for example, the proteins necessary for maturation of the human HIV virus. Once the three-dimensional structure of a protein is known or predicted, its function, as well as potential binding sites for drugs that inhibit its function, become tractable questions. The medical ramifications of the burgeoning results of protein structural biology, from gene replacement therapy to "rational" drug design, are well recognized by researchers in biomedical areas, and by a significant proportion of the general population. The purpose of this book is to introduce biomedical scientists to important areas of protein structural biology, and to provide an insightful orientation to the primary literature that shapes the field in each subject.

The chapters in this volume cover aspects of protein structural biology which have led to the recognition of fundamental relationships between protein structure and function.

Both eukaryotic and prokaryotic cells depend strongly on the function of ion pumps present in their membranes. The term ion pump, synonymous with active ion-transport system, refers to a membrane-associated protein that translocates ions uphill against an electrochemical potential gradient. Primary ion pumps utilize energy derived from chemical reactions or from the absorption of light, while secondary ion pumps derive the energy for uphill movement of one ionic species from the downhill movement of another species.

In the present volume, various aspects of ion pump structure, mechanism, and regulation are treated using mostly the ion-transporting ATPases as examples. One chapter has been devoted to a secondary ion pump, the Na+-Ca2+ exchanger, not only because of the vital role played by this transport system in regulation of cardiac contractility, but also because it exemplifies the interesting mechanistic and structural similarities between primary and secondary pumps.

The rapid expansion of the area of free radical biology in the last 25 years has occurred within a framework of assumptions and preconceived notions that has at times directed the course of this movement. The most dominant of these notions has been the view that free radical production is without exception a bad thing, and that the more efficient our elimination of these toxic substances, the better off we will be. The very important observation by Bernard Babior and colleagues in 1973 that activated phagocytes produce superoxide in order to kill micro organisms, served to illustrate that constructive roles are possible for free radicals. For many in the field, however, this merely underscored the deadly nature of oxygen-derived radicals, both from the microbe's point of view and from the host's as well. (Phagocyte-produced superoxide is responsible in part for the tissue injury manifested as inflammation. See Harris and Granger, Chapter 5, and Leff, Hybertson and Repine, Chapter 6.)Mother Nature, however, has a penchant for being able to make a silk purse from a sow's ear. If one is dealt a bad hand, one must simply make the best of it. After two decades of focusing on the destructive side of free radicals, the last few years have begun to reveal a new and finer perspective on free radical metabolism - a role in regulation of cellular function (see Schulze-Osthoff and Baeuerle, Chapter 2). Evidence from a number of sources suggests that an increase in the oxidative status of cell encourages that cell to grow and divide. Increasing the expression of mangnese superoxide dismutase can suppress the malignant phenotype of melanon cells (see Oberley and Oberley, Chapter 3). Oxidative stress beyond a certain poitosis (from the Greek, literally "to fall apart"). Is this suicide response an evolutionary fail-safe device to curtail tumorogenesis? Does oxidative stress-induced apoptosis account for the loss of immune cells in AIDS (see Flores and McCor Chapter 4)?This volume attempts to present the spectrum of roles, both good and bad played by active oxygen species as understood at this point in the evolution of this field of free radical biology.

This volume brings together a set of reviews that provide a summary of our current knowledge of the proteolytic machinery and of the pathways of protein breakdown of prokaryotic and eukaryotic cells. Intracellular protein degradation is much more than just a mechanism for the removal of incorrectly folded or damaged proteins. Since many short-lived proteins have important regulatory functions, proteolysis makes a significant contribution to many cellular processes including cell cycle regulation and transciptional control. In addition, limited proteolytic cleavage can provide a rapid and efficient mechanism of enzyme activation or inactivation in eukaryotic cells.In the first chapter, Maurizi provides an introduction to intracellular protein degradation, describes the structure and functions of bacterial ATP-dependent proteases, and explores the relationship between chaperone functions and protein degradation. Many of the principles also apply to eukaryotic cells, although the proteases involved are often not the same. Interestingly, homologues of one of the bacterial proteases, Ion protease, have been found in mitochondria in yeast and mammals, and homologues of proteasomes, which are found in all eukaryotic cells (see below), have been discovered in some eubacteria. Studies of proteolysis in yeast have contributed greatly to the elucidation of both lysosomal (vacuolar) and nonlysosomal proteolytic pathways in eukaryotic cells. Thumm and Wolf (chapter 2) describe studies that have elucidated the functions of proteasomes in nonlysosomal proteolysis and the contributions of lysosomal proteases to intracellular protein breakdown. Proteins can be selected for degradation by a variety of differen mechanisms. The ubiquitin system is one complex and highly regulated mechanism by which eukaryotic proteins are targetted for degradation by proteosomes. In chapter 3, Wilkinson reviews the components and functions of the ubiquitin system and considers some of the known substrates for this pathway which include cell cycle and transcriptional regulators. The structure and functions of proteosomes and their regulatory components are described in the two subsequent chapters by Tanaka and Tanahashi and by Dubiel and Rechsteiner. Proteasomes were the first known example of threonine proteases. They are multisubunit complexes that, in addition to being responsible for the turnover of most short-lived nuclear and cytoplasmic protein, are also involved in antigen processing for presentation by the MHC class I pathway. Recent studies reviewed by McCracken and colleagues (chapter 6) lead to the exciting conclusion that some ER-associated proteins are degraded by cytosolic proteasomes. Lysosomes are responsible for the degradation of long-lived proteins and for the enhanced protein degradation observed under starvation conditions. In chapter 7 Knecht and colleagues review the lysosomal proteases and describe studies of the roles of lysosomes and the mechanisms for protein uptake into lysosomes. Methods of measuring the relative contribution of different proteolytic systems (e.g., ubiquitin-proteasome pathway, calcium-dependent proteases, lysosomes) to muscle protein degradation, and the conclusions from such studies, are reviewed by Attai and Taillinder in the following chapter. Finally, proteases play an important role in signaling apoptosis by catalyzing the limited cleavage of enzymes. Mason and Beyette review the role of the major players, caspases, which are both activated by and catalyze limite proteolysis, and also consider the involvement of other protoelytic enzymes in this pathway leading cell death.

The aim of "The Adhesive Interaction of Cells" has been to assemble a series of reviews by leading international experts embracing many of the most important recent developments in this rapidly expanding field. The purpose of all biological research is to understand the form and function of living organisms and, by comprehending the normal, to find explanations and remedies for the abnormal and for disease conditions. The molecules involved in cell adhesion are of fundamental importance to the structure and function of all multicellular organisms. In this book, the contributors focus on the systems of vertebrates, especially mammals, since these are most relevant to human disease. It would have been equally possible to concentrate on developmental processes and adhesion in lower organisms. A major function of adhesion molecules is to bind cells to each other or to the extracellular matrix, but they are much more than "glue". Adhesions in animal tissues must be dynamic-forming, persisting, or declining in regulated fashion- to facilitate the mobility and turnover of tissue cells. Moreover, the majority of adhesion molecules are transmembrane molecules and thus provide links between the cells and their surroundings. This gives rise to another major function of adhesion molecules, the capacity to transduce signals across the hydrophobic barrier imposed by the plasma membrane. Such signal transduction is crucially important to many aspects of cellular function including the regulation of cell motility, gene expression, and differentiation. The work in this book progresses through four sections. Part I discusses the four major families of adhesion molecules themselves, the integrins (Green and Humphries), the cadherins (Stappert and Kemler), the selectins (Tedder et al.) and the immunoglobulin superfamily (Simmons); part 2 considers junctional complexes involved in cell interactions: focal adhesions and adherens junctions (Ben Ze'ev), desmosomes (Garrod et al.), and tight junctions (Citi and Cordenonsi). The signaling role of adhesion molecules is the focus of part 3, through integrins and the extracellular matrix (Edwards and Streuli), through platelet adhesion (Du and Ginsberg), and in the nervous system (Hemperley). In part 4, the aim is to show how adhesive phenomena contribute to important aspects of cell behavior and human health. Leukocyte trafficking (Haskard et al.), cancer metastasis (Marshall and Hart), cell migration (Paleck et al.), and implantation and placentation (Damsky et al.) are the topics considered in depth.The different sections are, of course, not mutually exclusive: it is both undesirable and impossible to separate structure from function when considering cell adhesion. Each chapter has its unique features, but some overlap is both invevitable and valuable since it provides different perspectives on closely related topics. We hope that the whole contributes a valuable and stimulating consideration of this important topic.

As a result of the key advances made more than 30 years ago, specifically the ability to isolate islets of Langerhans from the pancreas, the ability to measure insulin accurately by immunoasay, and the development of microchemical techniques for studying cells and their components, many research volumes, symposium reports, and original papers have been produced. This explosion of interest has probably had at least three stimuli:

1. the inherent scientific interest in understanding secretion of the pancreatic �-cell2. the �-cells relevance to a very common disease3. the availability of funding from specific sources related to diabetes research, for instance, Juvenile Diabetes Foundation International and the British Diabetic Association.

As a result of all this activity, detailed scientific literature including research reviews are readily available.

Surprisingly enough, there are relatively few attempts to summarize this great bulk of knowledge in a way that is accessible to the newcomer to this field and this book is intended to bridge this gap.

The objective in editing this volume was twofold: to provide a reasoned overview of the field as well as to furnish one that provided this overview within the context of the intellectual boundaries of those who initially attempted to define the purview of gap junction research. The latter objective has been realized by selecting the topics for review in this volume. The former objective was achieved by securing the cooperation of leaders in their fields as chapter co-authors.

After a decade of dominance by recombinant DNA technology, the field of molecular and cell biology is witnessing a renewed interest in techniques and approaches that are not driven by DNA acrobatics. In hindsight, this is an inevitable outcome. Deoxyribonucleic acid is not the master; it is only a storage house. If one wishes to know how cells work, the secret is not to be found in DNA, but rather in everything outside DNA. Science based on DNA is useful but does not itself solve the problem. It is most fortunate that at the height of the DNA phenomenon, there remain scientists who continue to probe cells by non-DNA means. Suddenly, people with such expertise are in high demand.

In this volume, some truly original scientists take the time to tell us their stories of innovations-some almost iconoclastic. All of these researchers have pioneered approaches that were long neglected; moreover, each is now in the fruitful phase of great harvests. It is a wonderful lesson for graduate students and postdoctorates that, although not being in the pack might be risky, the reward of such work is sweeter. As in physics, biology needs more young people who think like Richard Feynman - the ultimate iconoclast.

On the surface, the eight chapters of this volume appear to be diverse, but they are not. If our purpose is to understand cells, we must stop the habit of constantly dissecting leaves. Once in a while we have to see which forest we are in. The contributions included herein cannot cover the whole cell, but they give a sufficient flavor to arouse a desire to think more globally about cells. At a time when our field is in danger of being buried by thousands of kinases and phosphatases, these chapters inform our intended audiences that there are other ways - other techniques, other approaches, other thinkings, and other stories. We need all of them to appreciate the holistic aspect of cells, which has been a taboo until now.

Research into the basic mechanisms of photosynthesis has a long and distinguished history and has consistently been at the forefront of science. The success of this research, particularly in recent years, suggests that photosynthesis may turn out to be the first complex biological system to have its structure, function, and regulation described in rigorous chemical terms at the atomic level. It is likely that such knowledge will help us to tackle perhaps the most vital problem facing mankind, namely our need for a continuous and nonpolluting source of energy. The benefit may come by providing a "blueprint"for new technologies able to carry out efficient conversion of solar energy based on the principles of biological systems, and/or creating highly efficient "energy crops" sufficiently hardy to grow in a wide range of environments. The former is likely to involve new developments in material sciences while the latter will call on the rapidly advancing techniques of genetic engineering. The contents of this volume review some of the most important developments which are a part of the drive towards the overall goal of obtaining the complete description of the photosynthetic processes at the molecular level. The topics covered have been carefully selected and represent the wide spectrum of the subject.

It is now clear from a wide range of research that cytoplasm is not merely a buffered solution of proteins and enzymes but contains a series of complex filamentous structures. The cytoskeleton is the collective term given to these filaments. There is a considerable amount of data available on the protein composition of the major filament systems (microfilaments, microtubules, and intermediate filaments) but we are still comparatively ignorant about the role of the cytoskeleton in cell physiology. However such major cytoplasmic components (actin and tubulin, the monomeric constituents of microfilaments and microtubules, are major cell proteins) must have important roles to play in cell function, and investigations into the functional role of the cytoskeleton currently represent a major area of cell biological research.In recent years rapid advances in molecular biology have begun to influence research on the cytoskeleton. This trend is sure to continue and the techniques of molecular biology and genetics are set to make major contributions to our understanding of the cytoskeleton, as illustrated in this volume by several reviews; the use of transfection techniques by Ben-Ze'ev, the power of Drosophila genetics is described by Fyrberg and the major advances made in the inesin field using molecular approaches as described by Cyr et al. The chapters by Fyrberg and Cyr et al. also illustrate two other areas where major advances in our understanding of the cytoskeleton is occuring; the great array of different motor proteins involved in intracellular movements and the study of the cytoskeleton in developmental biology. Overall, Volume 12 in this series illuminates our increasing knowledge of the important roles of the cytoskeleton in cell function, particularly how it is central to metabolic organization, intracellular transport, interactions with matrix, and nerve function. Our knowledge of the cytoskeleton is now reaching a stage where it is clear that abnormalities in the organization of the cytoskeleton can lead to important clinical manifestations of disease; an example of how such research is now impinging on medical science is presented in the final chapter by Lane on keratin diseases.