Annual Review of Microbiology - Volume 61, 2007

I have dedicated the past 46 years of my life to science and I expect to be active in research for many more years. I have been lucky in my professional life. During my postdoctoral years I discovered two proteins that I showed to be involved in the initiation of protein synthesis. Working with bacteriophage ø29 for the past 40 years, we have made many interesting findings. Among them is the discovery of a protein covalently linked to the 5′ ends of ø29 DNA that we later showed to be the primer for the initiation of ø29 DNA replication. Also, the finding of the ø29 DNA polymerase with its properties of high processivity, strand displacement, and high fidelity has been very rewarding. The ø29 DNA polymerase has become the ideal enzyme for DNA amplification, both rolling circle and whole-genome amplification. I also am happy because I have worked with many brilliant students and collaborators over the years, most of whom have become excellent scientists.

Microbial communities contain unparalleled complexity, making them difficult to describe and compare. Characterizing this complexity will contribute to understanding the ecological processes that drive microbe-host interactions, bioremediation, and biogeochemistry. Moreover, an estimate of species richness will provide an indication of the completeness of a community profile. Such estimates are difficult, however, because community structure rarely fits a well-defined distribution. We present a model based on the word usage in books to illustrate the power of statistical tools in describing microbial communities and suggesting biological hypotheses. The model also generates data to test these methods when there are insufficient data in the literature. For example, by simulating the word distribution in books, we can predict the number of words that must be read to estimate the size of the vocabulary used to write the book. Combined with other models that have been used to make inaccessible problems tractable, our book model offers a unique approach to the complex problem of describing microbial diversity.

Isoniazid (INH) is one of the most efficient drugs for the treatment of Mycobacterium tuberculosis infections. Despite its rather simple chemical structure, the mechanism by which INH kills M. tuberculosis is complex. A full understanding of the mechanisms of action of INH required the development of genetic tools in M. tuberculosis. Herein, we discuss the different hypotheses that have been used to describe INH action against M. tuberculosis over the past 50 years in terms of the pregenetic and genetic era. We also review the different mechanisms of INH resistance and propose what we think is the means by which INH kills M. tuberculosis.

Production of industrial aromatic chemicals from renewable resources could provide a competitive alternative to traditional chemical synthesis routes. This review describes the engineering of microorganisms for the production of p-hydroxycinnamic acid (pHCA) and p-hydroxystyrene (pHS) from glucose. The initial process concept was demonstrated using a tyrosine-producing Escherichia coli strain that overexpressed both fungal phenylalanine/tyrosine ammonia lyase (PAL) and bacterial pHCA decarboxylase (pdc) genes. Further development of this bioprocess resulted in uncoupling the pHCA and pHS production steps to mitigate their toxicity to the production host. The final process consists of a fermentation step to convert glucose to tyrosine using a tyrosine-overproducing E. coli strain. This step is followed by a single biotransformation reaction to deaminate tyrosine to pHCA through immobilized E. coli cells that overexpress the Rhodotorula glutinis PAL gene. Finally, chemical decarboxylation of pHCA produces pHS. This multifaceted approach, which integrates biology, chemistry, and engineering, has allowed development of an economical process at scales suitable for industrial applications.

The RNA degradosome of Escherichia coli is a multiprotein complex involved in the degradation of mRNA. The principal components are RNase E, PNPase, RhlB, and enolase. RNase E is a large multidomain protein with an N-terminal catalytic region and a C-terminal noncatalytic region that is mostly natively unstructured protein. The noncatalytic region contains sites for binding RNA and for protein-protein interactions with other components of the RNA degradosome. Several recent studies suggest that there are alternative forms of the RNA degradosome depending on growth conditions or other factors. These alternative forms appear to modulate RNase E activity in the degradation of mRNA. RNA degradosome-like complexes appear to be conserved throughout the Proteobacteria, but there is a surprising variability in composition that might contribute to the adaptation of these bacteria to the enormously wide variety of niches in which they live.

Autotransporters are a large and diverse superfamily of proteins produced by pathogenic gram-negative bacteria that are composed of an N-terminal passenger domain, which typically harbors a virulence function, and a C-terminal β domain. It has long been known that the β domain anchors the protein to the outer membrane and facilitates transport of the passenger domain into the extracellular space. Despite the apparent simplicity of the autotransporter pathway, several aspects of autotransporter biogenesis remain poorly understood, most notably the mechanism by which the passenger domain is translocated across the outer membrane. Here we review recent evidence that the enormous sequence diversity of both passenger and β domains belies a remarkable conservation of structure. We also discuss insights into each stage of autotransporter biogenesis that have emerged from recent structural, biochemical, and imaging studies.

The use of photochemical reaction centers to convert light energy into chemical energy, chlorophototrophy, occurs in organisms belonging to only five eubacterial phyla: Cyanobacteria, Proteobacteria, Chlorobi, Chloroflexi, and Firmicutes. All chlorophototrophs synthesize two types of pigments: (a) chlorophylls and bacteriochlorophylls, which function in both light harvesting and uniquely in photochemistry; and (b) carotenoids, which function primarily as photoprotective pigments but can also participate in light harvesting. Although hundreds of carotenoids have been identified, only 12 types of chlorophylls (Chl a, b, d; divinyl-Chl a and b; and 8¹-hydroxy-Chl a) and bacteriochlorophylls (BChl a, b, c, d, e, and g) are currently known to occur in bacteria. This review summarizes recent progress in the identification of genes and enzymes in the biosynthetic pathways leading to Chls and BChls, the essential tetrapyrrole cofactors of photosynthesis, and addresses the mechanisms for generating functional diversity for solar energy capture and conversion in chlorophototrophs.

Cyclic diguanylate (c-di-GMP) is a bacterial second messenger of growing recognition involved in the regulation of a number of complex physiological processes. This review describes the biosynthesis and hydrolysis of c-di-GMP and several mechanisms of regulation of c-di-GMP metabolism. The contribution of c-di-GMP to regulating biofilm formation and motility, processes that affect pathogenesis of many bacteria, is described, as is c-di-GMP regulation of virulence gene expression. Finally, ways in which c-di-GMP may mediate these regulatory effects are proposed.

Virus replication and virus assembly often occur in virus inclusions or virus factories that form at pericentriolar sites close to the microtubule organizing center or in specialized nuclear domains called ND10/PML bodies. Similar inclusions called aggresomes form in response to protein aggregation. Protein aggregates are toxic to cells and are transported along microtubules to aggresomes for immobilization and subsequent degradation by proteasomes and/or autophagy. The similarity between aggresomes and virus inclusions raises the possibility that viruses use aggresome pathways to concentrate cellular and viral proteins to facilitate replication and assembly. Alternatively, aggresomes may be part of an innate cellular response that recognizes virus components as foreign or misfolded and targets them for storage and degradation. Insights into the possible roles played by aggresomes during virus assembly are emerging from an understanding of how virus inclusions form and how viral proteins are targeted to them.

The gut of the earthworm constitutes a mobile anoxic microzone to which the microorganisms of aerated soils are subjected. During gut passage, the in situ factors of the earthworm gut, which include anoxia and high concentrations of organic substrates, appear to greatly stimulate a subset of ingested soil microorganisms, including denitrifying and fermentative bacteria. The selective stimulation of ingested soil microbes by the unique microconditions of the earthworm gut (a) results in the in vivo emission of denitrification-derived dinitrogen (N2) and the greenhouse gas nitrous oxide (N2O) by the earthworm, and (b) might affect the fitness, culturability, and diversity of certain members of soil microbial biomes. These observations illustrate the impact that soil macrofauna might have on terrestrial nitrogen cycle processes via their transient hosting of ingested prokaryotes.

The cell envelope of gram-negative bacteria consists of two membranes, the inner and the outer membrane, that are separated by the periplasm. The outer membrane consists of phospholipids, lipopolysaccharides, integral membrane proteins, and lipoproteins. These components are synthesized in the cytoplasm or at the inner leaflet of the inner membrane and have to be transported across the inner membrane and through the periplasm to assemble eventually in the correct membrane. Recent studies in Neisseria meningitidis and Escherichia coli have led to the identification of several machineries implicated in these transport and assembly processes.

One of the strongest and most noticeable responses of Bacillus subtilis cells to a range of stress and starvation stimuli is the dramatic induction of about 150 SigB-dependent general stress genes. The activity of SigB itself is tightly regulated by a complex signal transduction cascade with at least three main signaling pathways that respond to environmental stress, energy depletion, or low temperature. The SigB-dependent response is conserved in related gram-positive bacteria but is missing in strictly anaerobic or in some facultatively anaerobic gram-positive bacteria. It covers functions from nonspecific and multiple stress resistance to the control of virulence in pathogenic bacteria. A comprehensive understanding of this crucial stress response is essential not only for bacterial physiology but also for applied microbiology, including pathogenicity and pathogen control.

The shewanellae are aquatic microorganisms with worldwide distribution. Their hallmark features include unparalleled respiratory diversity and the capacity to thrive at low temperatures. As a genus the shewanellae are physiologically diverse, and this review provides an overview of the varied roles they serve in the environment and describes what is known about how they might survive in such extreme and harsh environments. In light of their fascinating physiology, these organisms have several biotechnological uses, from bioremediation of chlorinated compounds, radionuclides, and other environmental pollutants to energy-generating biocatalysis. The ecology and biotechnology of these organisms are intertwined, with genomics playing a key role in our understanding of their physiology.

In eukaryotic cells, repair of DNA double-strand breaks (DSBs) by the nonhomologous end-joining (NHEJ) pathway is critical for genomic stability. A functionally homologous repair apparatus, composed of Ku and a multifunctional DNA ligase (LigD), has recently been identified in many prokaryotes. Eukaryotic organisms employ a large number of factors to repair breaks by NHEJ. In contrast, the bacterial NHEJ complex is a two-component system that, despite its relative simplicity, possesses all of the break-recognition, end-processing, and ligation activities required to facilitate the complex task of DSB repair. Here, we review recent discoveries on the structure and function of the bacterial NHEJ repair apparatus. In particular, we discuss the evolutionary origins of this DSB repair pathway, how the diverse activities within the prokaryotic end-joining complex cooperate to facilitate DSB repair, the physiological roles of bacterial NHEJ, and finally, the essential function of NHEJ in the life cycle of mycobacteriophage.

This review describes some of the recent highlights taken from the studies of Rhodobacter sphaeroides 2.4.1. The review is not intended to be comprehensive, but to reflect the bias of the authors as to how the availability of a sequenced and annotated genome, a gene-chip, and proteomic profile as well as comparative genomic analyses can direct the progress of future research in this system.

Bacterial cells contain many large, spatially extended assemblies of ions, molecules, and macromolecules, called hyperstructures, that are implicated in functions that range from DNA replication and cell division to chemotaxis and secretion. Interactions between these hyperstructures would create a level of organization intermediate between macromolecules and the cell itself. To explore this level, a taxonomy is needed. Here, we describe classification criteria based on the form of the hyperstructure and on the processes responsible for this form. These processes include those dependent on coupled transcription-translation, protein-protein affinities, chromosome site-binding by protein, and membrane structures. Various combinations of processes determine the formation, maturation, and demise of many hyperstructures that therefore follow a trajectory within the space of classification by form/process. Hence a taxonomy by trajectory may be desirable. Finally, we suggest that working toward a taxonomy based on speculative interactions between hyperstructures promises most insight into life at this level.

The endolithic environment, the pore space in rocks, is a ubiquitous microbial habitat and an interface between biology and geology. Photosynthesis-based endolithic communities inhabit the outer centimeters of rocks exposed to the surface, and offer model systems for microbial ecology, geobiology, and astrobiology. Endolithic ecosystems are among the simplest microbial ecosystems known and as such provide tractable models for testing ecological hypotheses. Such hypotheses have been difficult to test because microbial ecosystems are extraordinarily diverse. We review here recent culture-independent, ribosomal RNA-based studies that evaluate hypotheses about endolithic ecosystems, and provide insight for understanding general principles in microbial ecology. Comparison of endolithic communities supports the principle that patterns of microbial diversity are governed by similar principles observed in macroecological systems. Recent results also explore geobiological processes that shape the current biosphere and potentially provide clues to life's history on Earth and where to seek life elsewhere in the Solar System.

A wide range of Bacteria and Archaea sense cellular 2-oxoglutarate (2OG) as an indicator of nitrogen limitation. 2OG sensor proteins are varied, but most of those studied belong to the PII superfamily. Within the PII superfamily, GlnB and GlnK represent a widespread family of homotrimeric proteins (GlnB-K) that bind and respond to 2OG and ATP. In some bacterial phyla, GlnB-K proteins are covalently modified, depending on enzymes that sense cellular glutamine as an indicator of nitrogen sufficiency. GlnB-K proteins are central clearing houses of nitrogen information and bind and modulate a variety of nitrogen assimilation regulators and enzymes. NifI1 and NifI2 comprise a second widespread family of PII proteins (NifI) that are heteromultimeric, respond to 2OG and ATP, and bind and regulate dinitrogenase in Euryarchaeota and many Bacteria.

The field of bacterial phosphorus (P) metabolism has undergone a significant transformation in the past decade owing to the elucidation of widespread and diverse pathways for the metabolism of reduced P compounds. The characterization of these pathways dramatically changes the current and narrow view of P metabolism and our understanding of the forms in which P is produced and available in the environment. In this review, recent investigations into the biochemical pathways and molecular genetics of reduced P metabolism in bacteria are discussed. Particular attention is paid to recently elucidated metabolic reactions and the genetic characterization of biosynthesis of organic reduced P compounds and to the pathways for oxidation of the inorganic reduced P compounds hypophosphite and phosphite.

Plants support a diverse array of bacteria, including parasites, mutualists, and commensals on and around their roots, in the vasculature, and on aerial tissues. These microbes have a profound influence on plant health and productivity. Bacteria physically interact with surfaces to form complex multicellular and often multispecies assemblies, including biofilms and smaller aggregates. There is growing appreciation that the intensity, duration, and outcome of plant-microbe interactions are significantly influenced by the conformation of adherent microbial populations. Biofilms on different tissues have unique properties, reflecting the prevailing conditions at those sites. Attachment is required for biofilm formation, and bacteria interact with plant tissues through adhesins including polysaccharides and surface proteins, with initial contact often mediated by active motility. Recognition between lectins and their cognate carbohydrates is a common means of specificity. Biofilm development and the resulting intimate interactions with plants often require cell-cell communication between colonizing bacteria.

Filamentous fungi are multicellular eukaryotic organisms known for nutrient recycling as well as for antibiotic and food production. This group of organisms also contains the most devastating plant pathogens and several important human pathogens. Since the first report of heterotrimeric G proteins in filamentous fungi in 1993, it has been demonstrated that G proteins are essential for growth, asexual and sexual development, and virulence in both animal and plant pathogenic filamentous species. Numerous G protein subunit and G protein–coupled receptor genes have been identified, many from whole-genome sequences. Several regulatory pathways have now been delineated, including those for nutrient sensing, pheromone response and mating, and pathogenesis. This review provides a comparative analysis of G protein pathways in several filamentous species, with discussion of both unifying themes and important unique signaling paradigms.

Data from protist genomes suggest that eukaryotes show enormous variability in their gene complements, especially of genes coding regulatory proteins. Overall counts of eukaryotic signaling proteins show weak nonlinear scaling with proteome size, but individual superfamilies of signaling domains might show vast expansions in certain protists. Alteration of domain architectural complexity of signaling proteins and repeated lineage-specific reshaping of architectures might have played a major role in the emergence of new signaling interactions in different eukaryotes. Lateral transfer of various signaling domains from bacteria or from hosts, in parasites such as apicomplexans, appears to also have played a major role in the origin of new functional networks. Lineage-specific expansion of regulatory proteins, particularly of transcription factors, has played a critical role in the adaptive radiation of different protist lineages. Comparative genomics allows objective reconstruction of the ancestral conditions and subsequent diversification of several regulatory systems involved in phosphorylation, cyclic nucleotide signaling, Ubiquitin conjugation, chromatin remodeling, and posttranscriptional gene silencing.

The current need for antibiotics with novel target molecules has coincided with advances in technical approaches for the structural and functional analysis of the lantibiotics, which are ribosomally synthesized peptides produced by gram-positive bacteria. These peptides have antibiotic or morphogenetic activity and are structurally defined by the presence of unusual amino acids introduced by posttranslational modification. Lantibiotics are complex polycyclic molecules formed by the dehydration of select Ser and Thr residues and the intramolecular addition of Cys thiols to the resulting unsaturated amino acids to form lanthionine and methyllanthionine bridges, respectively. Importantly, the structural and functional diversity of the lantibiotics is much broader than previously imagined. Here we discuss this growing collection of molecules and introduce some recently discovered peptides, review advances in enzymology and protein engineering, and discuss the regulatory networks that govern the synthesis of the lantibiotics by the producing organisms.

The availability of whole-genome sequences for ammonia-oxidizing bacteria (AOB) has led to dramatic increases in our understanding of these environmentally important microorganisms. Their genomes are smaller than many other members of the proteobacteria and may indicate genome reductions consistent with their limited lifestyle. The genomes have a surprising level of gene repetition including genes for ammonia catabolism, iron acquisition, and insertion sequences. The gene profiles reveal limited genes for catabolism and transport of complex organic compounds, but complete pathways for some other compounds. This led to the observation of chemolithoheterotrophic growth of Nitrosomonas europaea. Genes for sucrose synthesis/degradation were identified. The core metabolic module of aerobic ammonia oxidation, the extraction of electrons from hydroxylamine to generate proton-motive force and reductant, has evolutionary roots in the denitrification inventory of anaerobic sulfur-dependent bacteria. The extension by ammonia monooxygenase provides a mechanism to feed this module using ammonia and O2.

Candida albicans is termed a dimorphic fungus because it proliferates in either a yeast form or a hyphal form. The switch between these forms is the result of a complex interplay of external and internal factors and is coordinated in part by polarity-regulating proteins that are conserved among eukaryotic cells. However, yeast and hyphal cells are not the only morphological states of C. albicans. The opaque form required for mating, the pseudohyphal cell, and the chlamydospore represent distinct cell types that form in response to specific genetic or environmental conditions. In addition, hyperextended buds can form as a result of various cell cycle–related stresses. Recent studies are beginning to shed light on some of the molecular controls regulating the various morphogenetic forms of this fascinating human pathogen.

Endospores formed by Bacillus, Clostridia, and related genera are encased in a protein shell called the coat. In many species, including B. subtilis, the coat is the outermost spore structure, and in other species, such as the pathogenic organisms B. anthracis and B. cereus, the spore is encased in an additional layer called the exosporium. Both the coat and the exosporium have roles in protection of the spore and in its environmental interactions. Assembly of both structures is a function of the mother cell, one of two cellular compartments of the developing sporangium. Studies in B. subtilis have revealed that the timing of coat protein production, the guiding role of a small group of morphogenetic proteins, and several types of posttranslational modifications are essential for the fidelity of the assembly process. Assembly of the exosporium requires a set of novel proteins as well as homologues of proteins found in the outermost layers of the coat and of some of the coat morphogenetic factors, suggesting that the exosporium is a more specialized structure of a multifunctional coat. These and other insights into the molecular details of spore surface morphogenesis provide avenues for exploitation of the spore surface layers in applications for biotechnology and medicine.

All cytoskeletal elements known from eukaryotic cells are also present in bacteria, where they perform vital tasks in many aspects of the physiology of the cell. Bacterial tubulin (FtsZ), actin (MreB), and intermediate filament (IF) proteins are key elements in cell division, chromosome and plasmid segregation, and maintenance of proper cell shape, as well as in maintenance of cell polarity and assembly of intracellular organelle-like structures. Although similar tasks are performed by eukaryotic cytoskeletal elements, the individual functions of FtsZ, MreBs, and IFs are different from those performed by their eukaryotic orthologs, revealing a striking evolutional plasticity of cytoskeletal proteins. However, similar to the functions of their eukaryotic counterparts, the functions conferred by bacterial cytoskeletal proteins are driven by their ability to form dynamic filamentous structures. Therefore, the cytoskeleton was a prokaryotic invention, and additional bacteria-specific cytoskeletal elements, such as fibril and MinD-type ATPases, that confer various functions in cell morphology and during the cell cycle have been observed in prokaryotes. The investigation of these elements will give fundamental information for all types of cells and can reveal the molecular mode of action of cytoskeletal, filament-forming proteins.