Genetic approaches to auxin action

Answers to long-standing questions concerning the molecular mechanism of auxin action and auxin's exact functions in plant growth and development are beginning to be uncovered through studies using mutant and transgenic plants. We review recent work in this area in vascular plants. A number of conclusions can be drawn from these studies. First, auxin appears essential for cell division and viability, as auxin auxotrophs isolated in tissue culture are dependent on auxin for growth and cannot be regenerated into plants even when auxin is supplied exogenously. Secondly, plants with transgenes that alter auxin levels are able to regulate cellular auxin concentrations by synthesis and conjugation; wild-type plants are probably also capable of such regulation. Thirdly, the phenotypes of transgenic plants with altered auxin levels and of mutant plants with altered sensitivity to auxin confirm earlier physiological studies which indicated a role for auxin in regulation of apical dominance, in development of roots and vascular tissue, and in the gravitropic response. Finally, the cloning of a mutationally identified gene important for auxin action, along with accumulating biochemical evidence, hints at a major role for protein degradation in the auxin response pathway.     

Approaches to understanding auxin action

The effects of auxin on plant growth and development have been studied for decades, but the molecular mechanisms of auxin action remain unknown. These mechanisms have primarily been investigated by characterization of auxin physiology mutants and analysis of auxin-binding proteins and auxin-regulated genes. These efforts are now converging, since some mutants have recently been shown to have altered expression of specific auxin-binding proteins and auxin-regulated genes. The features of these proteins and genes are providing the first tantalizing clues to the organization of auxin signal transduction pathways.     

Evolutionary patterns in auxin action

This review represents the first effort ever to survey the entire literature on auxin (indole-3-acetic acid, IAA) action in all plants, with special emphasis on the green plant lineage, including charophytes (the green alga group closest to the land plants), bryophytes (the most basal land plants), pteridophytes (vascular non-seed plants), and seed plants. What emerges from this survey is the surprising perspective that the physiological mechanisms for regulating IAA levels and many IAA-mediated responses found in seed plants are also present in charophytes and bryophytes, at least in nascent forms. For example, the available evidence suggests that the apical regions of both charophytes and liverworts synthesize IAA via a tryptophan-independent pathway, with IAA levels being regulated via the balance between the rates of IAA biosynthesis and IAA degradation. The apical regions of all the other land plants utilize the same class of biosynthetic pathway, but they have the potential to utilize IAA conjugation and conjugate hydrolysis reactions to achieve more precise spatial and temporal control of IAA levels. The thallus tips of charophytes exhibit saturable IAA influx and efflux carriers, which are apparently not sensitive to polar IAA transport inhibitors. By contrast, two divisions of bryophyte gametophytes and moss sporophytes are reported to carry out polar IAA transport, but these groups exhibit differing sensitivities to those inhibitors. Although the IAA regulation of charophyte development has received almost no research attention, the bryophytes manifest a wide range of developmental responses, including tropisms, apical dominance, and rhizoid initiation, which are subject to IAA regulation that resembles the hormonal control over corresponding responses in seed plants. In pteridophytes, IAA regulates root initiation and vascular tissue differentiation in a manner also very similar to its effects on those processes in seed plants. Thus, it is concluded that the seed plants did not evolve de novo mechanisms for mediating IAA responses, but have rather modified pre-existing mechanisms already operating in the early land plants. Finally, this paper discusses the encouraging prospects for investigating the molecular evolution of auxin action.     

Ion channels meet auxin action

The regulation of cell division and elongation in plants is accomplished by the action of different phytohormones. Auxin as one of these growth regulators is known to stimulate cell elongation growth in the aerial parts of the plant. Here, auxin enhances cell enlargement by increasing the extensibility of the cell wall and by facilitating the uptake of osmolytes such as potassium ions into the cell. Starting in the late 1990s, the auxin regulation of ion channels mediating K+ import into the cell has been studied in great detail. In this article we will focus on the molecular mechanisms underlying the modulation of K+ transport by auxin and present a model to explain how the regulation of K+ channels is involved in auxin-induced cell elongation growth.     

Genetic approaches to common diseases

Combined molecular and epidemiological studies are advancing our understanding of the genetic basis of multifactorial diseases. Several of the results obtained during the past year highlight methodological issues associated with these approaches. For example, the affected sib-pair method has been applied successfully to detect linkage between the angiotensinogen gene and susceptibility to hypertension, and a large multi-centre epidemiological study has demonstrated association of a polymorphism of the angiotensin-converting enzyme gene with increased risk of myocardial infarction. The study of Mendelian forms of multifactorial diseases has also led to many new results. These include the characterization of mutations in the glucokinase gene in maturity onset diabetes of the young, localization to chromosome 2 of a gene involved in familial colon cancer, and localization to chromosome 19 of a gene responsible for hemiplegic migraine. New insights have been provided into the genetics of multifactorial disorders such as diabetes and hypertension through the study of animal models. Localization of susceptibility loci in such models has recently led to the identification of new candidate genes that may be implicated in disease.     

Points of regulation for auxin action

There have been few examples of the application of our growing knowledge of hormone action to crop improvement. In this review we discuss what is known about the critical points regulating auxin action. We examine auxin metabolism, transport, perception and signalling and identify genes and proteins that might be keys to regulation, particularly the rate-limiting steps in various pathways. Certain mutants show that substrate flow in biosynthesis can be limiting. To date there is little information available on the genes and proteins of catabolism. There have been several auxin transport proteins and some elegant transport physiology described recently, and the potential for using transport proteins to manage free indole-3-acetic acid (IAA) concentrations is discussed. Free IAA is very mobile, and so while it may be more practical to control auxin action through managing the receptor and signalling pathways, the candidate genes and proteins through which this can be done remain largely unknown. From the available evidence, it is clear that the reason for so few commercial applications arising from the control of auxin action is that knowledge is still limited.     

Genetic approaches to studying peroxisome biogenesis

How proteins are imported into peroxisomes is a question attracting considerable interest at present. Peroxisomal proteins, including the integral membrane proteins of the membrane bounding the peroxisome, are synthesized on free cytoplasmic ribosomes. They assemble post-translationally into pre-existing peroxisomes. New peroxisomes are believed to form exclusively by division of old ones. Few molecular details of this process have been elucidated so far, but genetic approaches are now beginning to identify the proteins catalysing peroxisome assembly.     

Genetic approaches to memory storage.

The ability to remember is perhaps the most significant and distinctive feature of our mental life. We are who we are largely because of what we have learned and what we remember. In turn, impairments in learning and memory can lead to disorders that range from the moderately inconvenient benign senescent forgetfulness associated with normal aging to the devastating memory losses associated with Alzheimer disease. Of the various, higher-cognitive abilities that human beings possess, such as reasoning and language, memory is the only one that can be studied effectively in simple experimental organisms that are accessible to genetic manipulation, such as snails, flies and mice. In these organisms, the effectiveness of genetic approaches in the study of memory has improved significantly in the past five years. Below we review these advances.     

Genetic and genomic approaches to glomerulosclerosis

Chronic kidney disease (CKD) is common, progressive and expensive to manage. Although modifiable risk factors can be treated and outcomes improved, CKD remains a chronic disease with excessive morbidity and mortality. The completion of the human genome sequence and the advent of methodologies to define gene function provide new opportunities to manage and treat patients with CKD and other chronic diseases. Despite the lack of clear correspondence between genotype and phenotype and an obvious Mendelian inheritance pattern, CKD susceptibility has a genetic basis. In this review, we focus on recent studies of familial focal segmental glomerulosclerosis and the discoveries that have resulted from both genetic and genomic approaches used to understand its pathogenesis. Key slit diaphragm proteins were discovered using linkage analyses of these rare causes of glomerulosclerosis and subsequent work has characterized slit diaphragm function in health and disease. Podocyte dysfunction is now recognized as a key contributor to the functional and histologic derangements that characterize glomerular dysfunction in many common causes of CKD. In aggregate, these studies provide a paradigm for approaches to better define mechanisms of CKD and to identify novel therapeutic targets.     

Genetic approaches to harvesting lichen products

Lichens are symbiotic associations between fungi, green algae and/or cyanobacteria. They have a varied chemistry and produce many polyketide-derived compounds, including some, such as depsides and depsidones, that are rarely reported elsewhere. Although lichens have been appreciated in traditional medicines, their value has largely been ignored by the modern pharmaceutical industry because difficulties in establishing axenic cultures and conditions for rapid growth preclude their routine use in most conventional screening processes. Recently, molecular genetic techniques using PCR, genomic library construction and heterologous expression have provided an alternative approach to begin exploring the diversity of polyketide biosynthetic pathways in lichens. The techniques can be expanded to cover other pathway types and be integrated with conventional culture collection-based screening to provide a comprehensive search for novel chemical entities in these organisms.     

Genetic approaches to disease and regeneration

Cardiovascular disease is largely a consequence of coronary artery blockage through excessive proliferation of smooth muscle cells. It in turn leads to myocardial infarction and permanent and functionally devastating tissue damage to the heart wall. Our studies have revealed that elastin is a primary player in maintaining vascular smooth muscle cells in their dormant state and thus may be a useful therapeutic in vascular disease. By studying zebrafish, which unlike humans, can repair damage to heart muscle, we have begun to uncover some of the genes that seem necessary to undertake the de-differentiation steps that currently fail and prevent the formation of new proliferating cardiomyocytes at the site of damage in a mammalian heart.     

Genetic approaches to understanding sugar-response pathways

Plants as photoautotrophic organisms are able to produce the carbohydrates they require and have developed mechanisms to co-ordinate carbohydrate production and its metabolism. Carbohydrate-derived signals regulate the expression of genes involved in both photosynthesis and metabolism, and control carbohydrate partitioning. A number of genetic approaches have been initiated to understand sugar-response pathways in plants and identify the components involved. Screening strategies to date have been based on the effects of high sugar media on early seedling development or on changes in the enzyme activity or expression of sugar-responsive genes. These screens have established roles for plant hormones in sugar-response pathways, in particular for abscisic acid. The present emphasis on the role of plant hormones in sugar responses is due to the fact that mutants could be readily identified as belonging to these established pathways, but also results from the nature of the mutant screens in use. Progress is being made on the identification of mutants and genes that may be specific to sugar-signalling pathways. It is also expected that the modification of existing screens may target sugar-signalling pathways more directly. Genetic approaches may be especially useful in identifying components of novel signalling pathways unique to plants, and their combination with genomic and molecular approaches will guide future research.     

A molecular basis for auxin action.

The plant hormone auxin is central in the regulation of growth and development, however, the molecular basis for its action has remained enigmatic. In the absence of a molecular model, the wide range of responses elicited by auxin have been difficult to explain. Recent advances using molecular genetic approaches in Arabidopsis have led to the isolation of a number of key genes involved in auxin action. Of particular importance are genes involved in channelling polar auxin transport through the plant. In addition a model for auxin signal transduction, centred on regulated protein degradation, has been developed.