Annual Review of Genetics - Volume 37, 2003

Transposons are mobile genetic elements that can relocate from one genomic location to another. As well as modulating gene expression and contributing to genome plasticity and evolution, transposons are remarkably diverse molecular tools for both whole-genome and single-gene studies in bacteria, yeast, and other microorganisms. Efficient but simple in vitro transposition reactions now allow the mutational analysis of previously recalcitrant microorganisms. Transposon-based signature-tagged mutagenesis and genetic footprinting strategies have pinpointed essential genes and genes that are crucial for the infectivity of a variety of human and other pathogens. Individual proteins and protein complexes can be dissected by transposon-mediated scanning linker mutagenesis. These and other transposon-based approaches have reaffirmed the usefulness of these elements as simple yet highly effective mutagens for both functional genomic and proteomic studies of microorganisms.

Cells have high-fidelity polymerases whose task is to accurately replicate the genome, and low-fidelity polymerases with specialized functions. Although some of these low-fidelity polymerases are exceptional in their ability to replicate damaged DNA and restore the undamaged sequence, they are error prone on undamaged DNA. In fact, these error-prone polymerases are sometimes used in circumstances where the capacity to make errors has a selective advantage. The mutagenic potential of the error-prone polymerases requires that their expression, activity, and access to undamaged DNA templates be regulated. Here we review these specialized polymerases with an emphasis on their biological roles.

Differences in skin and hair color are principally genetically determined and are due to variation in the amount, type, and packaging of melanin polymers produced by melanocytes secreted into keratinocytes. Pigmentary phenotype is genetically complex and at a physiological level complicated. Genes determining a number of rare Mendelian disorders of pigmentation such as albinism have been identified, but only one gene, the melanocortin 1 receptor (MCR1), has so far been identified to explain variation in the normal population such as that leading to red hair, freckling, and sun-sensitivity. Genotype-phenotype relations of the MC1R are reviewed, as well as methods to improve the phenotypic assessment of human pigmentary status. It is argued that given advances in model systems, increases in technical facility, and the lower cost of genotype assessment, the lack of standardized phenotype assessment is now a major limit on advance.

Thiol-based regulatory switches play central roles in cellular responses to oxidative stress, nitrosative stress, and changes in the overall thiol-disulfide redox balance. Protein sulfhydryls offer a great deal of flexibility in the different types of modification they can undergo and the range of chemical signals they can perceive. For example, recent work on OhrR and OxyR has clearly established that disulfide bonds are not the only cysteine oxidation products that are likely to be relevant to redox sensing in vivo. Furthermore, different stresses can result in distinct modifications to the same protein; in OxyR it seems that distinct modifications can occur at the same cysteine, and in Yap1 a partner protein ensures that the disulfide bond induced by peroxide stress is different from the disulfide bond induced by other stresses. These kinds of discoveries have also led to the intriguing suggestion that different modifications to the same protein can create multiple activation states and thus deliver discrete regulatory outcomes. In this review, we highlight these issues, focusing on seven well-characterized microbial proteins controlled by thiol-based switches, each of which exhibits unique regulatory features.

Pseudogenes have been defined as nonfunctional sequences of genomic DNA originally derived from functional genes. It is therefore assumed that all pseudogene mutations are selectively neutral and have equal probability to become fixed in the population. Rather, pseudogenes that have been suitably investigated often exhibit functional roles, such as gene expression, gene regulation, generation of genetic (antibody, antigenic, and other) diversity. Pseudogenes are involved in gene conversion or recombination with functional genes. Pseudogenes exhibit evolutionary conservation of gene sequence, reduced nucleotide variability, excess synonymous over nonsynonymous nucleotide polymorphism, and other features that are expected in genes or DNA sequences that have functional roles. We first review the Drosophila literature and then extend the discussion to the various functional features identified in the pseudogenes of other organisms. A pseudogene that has arisen by duplication or retroposition may, at first, not be subject to natural selection if the source gene remains functional. Mutant alleles that incorporate new functions may, nevertheless, be favored by natural selection and will have enhanced probability of becoming fixed in the population. We agree with the proposal that pseudogenes be considered as potogenes, i.e., DNA sequences with a potentiality for becoming new genes.

Paramecium bursaria chlorella virus (PBCV-1) is the prototype of a family of large, icosahedral, plaque-forming, dsDNA viruses that replicate in certain unicellular, eukaryotic chlorella-like green algae. Its 330-kb genome contains ∼373 protein-encoding genes and 11 tRNA genes. The predicted gene products of ∼50% of these genes resemble proteins of known function, including many that are unexpected for a virus, e.g., ornithine decarboxylase, hyaluronan synthase, GDP-D-mannose 4,6 dehydratase, and a potassium ion channel protein. In addition to their large genome size, the chlorella viruses have other features that distinguish them from most viruses. These features include: (a) The viruses encode multiple DNA methyltransferases and DNA site-specific endonucleases. (b) The viruses encode at least some, if not all, of the enzymes required to glycosylate their proteins. (c) PBCV-1 has at least three types of introns, a self-splicing intron in a transcription factor-like gene, a spliceosomal processed intron in its DNA polymerase gene, and a small intron in one of its tRNA genes. (d) Many chlorella virus-encoded proteins are either the smallest or among the smallest proteins of their class. (e) Accumulating evidence indicates that the chlorella viruses have a very long evolutionary history.

The enzyme lactase that is located in the villus enterocytes of the small intestine is responsible for digestion of lactose in milk. Lactase activity is high and vital during infancy, but in most mammals, including most humans, lactase activity declines after the weaning phase. In other healthy humans, lactase activity persists at a high level throughout adult life, enabling them to digest lactose as adults. This dominantly inherited genetic trait is known as lactase persistence. The distribution of these different lactase phenotypes in human populations is highly variable and is controlled by a polymorphic element cis-acting to the lactase gene. A putative causal nucleotide change has been identified and occurs on the background of a very extended haplotype that is frequent in Northern Europeans, where lactase persistence is frequent. This single nucleotide polymorphism is located 14 kb upstream from the start of transcription of lactase in an intron of the adjacent gene MCM6. This change does not, however, explain all the variation in lactase expression.

The anterior-posterior axis of the Caenorhabditis elegans zygote forms shortly after fertilization when the sperm pronucleus and its associated centrosomal asters provide a cue that establishes the anterior-posterior (AP) body axis. In response to this cue, the microfilament cytoskeleton polarizes the distribution of a group of widely conserved, cortically localized regulators called the PAR proteins, which are required for the first mitotic division to be asymmetric. These asymmetries include a posterior displacement of the first mitotic spindle and the differential segregation of cell-fate determinants to the anterior and posterior daughters produced by the first cleavage of the zygote. Here we review recent advances in our understanding of the mechanisms that polarize the one-cell zygote to generate an AP axis of asymmetry.

The mitotic spindle segregates chromosomes to opposite ends of the cell in preparation for cell division. Chromosome attachment to the spindle is monitored by the spindle assembly checkpoint, and at least in yeast cells, penetration of one spindle pole into the bud is monitored by the spindle position checkpoint. We review the historical origins of these checkpoints and recent progress in understanding their surveillance pathways. We also highlight fascinating but as yet unresolved questions, and examine crosstalk between the checkpoints.

Lateral gene transfer (LGT) is now known to be a major force in the evolution of prokaryotic genomes. To date, most analyses have focused on either (a) verifying phylogenies of individual genes thought to have been transferred, or (b) estimating the fraction of individual genomes likely to have been introduced by transfer. Neither approach does justice to the ability of LGT to effect massive and complex transformations in basic biology. In some cases, such transformation will be manifested as the patchy distribution of a seemingly fundamental property (such as aerobiosis or nitrogen fixation) among the members of a group classically defined by the sharing of other properties (metabolic, morphological, or molecular, such as small subunit ribosomal RNA sequence). In other cases, the lineage of recipients so transformed may be seen to comprise a new group of high taxonomic rank (“class” or even “phylum”). Here we review evidence for an important role of LGT in the evolution of photosynthesis, aerobic respiration, nitrogen fixation, sulfate reduction, methylotrophy, isoprenoid biosynthesis, quorum sensing, flotation (gas vesicles), thermophily, and halophily. Sometimes transfer of complex gene clusters may have been involved, whereas other times separate exchanges of many genes must be invoked.

Research into the mechanisms underlying the process of aging is emerging as an exciting area of biomedical research. Observations challenging the fundamental assumptions of aging have begun to rejuvenate the field, opening up aging research to fresh ideas and approaches. Genetic approaches, which have been successfully used to understand other complex biological phenomena, are beginning to reveal important patterns and conservations between the processes of aging in a variety of species including yeast, nematodes, flies, and mice. A combination of candidate and random gene alteration approaches, particularly in the fruitfly model system, Drosophila melanogaster, should prove to be especially valuable for elucidating the primary physiological systems involved in aging and life span determination.

Sexual reproduction results from the fusion of gametes in which the chromatin configuration of maternal and paternal chromosomes is distinct at fertilization. Although many of the differences are erased during successive cellular divisions and chromatin modifications, some are retained in both somatic and germline cells. These epigenetic modifications can confer different characteristics on maternal and paternal chromosomes and such differences can be selected during any process that has the ability to distinguish between homologues. The end result of these selective forces are parental origin effects, writ large. The range of effects observed, including transcriptional imprinting and effects on chromosome segregation and heterochromatization, reflects the diversity of selective forces in operation. However, a closer look at these effects suggests that parental origin–dependent differences in chromatin structure might be subject to some common forces and that these forces may explain many of the “nontranscriptional” parental origin effects observed in mammals.

Vernalization is the process whereby the floral transition is promoted through exposure of plants to long periods of cold temperature or winter. A requirement for vernalization aligns flowering with the seasons to ensure that their reproductive phase occurs in favorable conditions. The mitotic stability of vernalization, suggestive of an epigenetic mechanism, has intrigued researchers for many years. Genetic analysis of the vernalization requirement in Arabidopsis has identified key floral repressor genes, FRI and FLC. The action of these floral repressors is antagonized by vernalization and the activity of a set of genes grouped into the autonomous floral pathway. Analysis of the vernalization pathway has defined a series of epigenetic regulators crucial for “cellular-memory” of the cold signal, whereas the autonomous pathway appears to function in part through posttranscriptional mechanisms. The mechanism of the vernalization requirement, which is now being explored in a range of plant species, should uncover the evolutionary origins of this key agronomic trait.

The most studied example of transmission ratio distortion (TRD) in mice is that of the t-complex. This is a variant region of Chromosome 17 which exists as a polymorphism in wild mice. Males heterozygous for a t-haplotype and a normal Chr 17 transmit the t-haplotype to >50% of their young, up to 99%. Homozygous males are sterile. The TRD produced by the t-complex is due to the action of three or more distorter genes (Tcd) on a responder gene (Tcr). t-Haplotypes are maintained intact by crossover suppression induced by four neighboring inversions, the Tcd and Tcr loci lying in different inversions. Sperm formation is normal in t/t males, but sperm function is impaired through gross defects in sperm motility. The responder gene has been identified as a fusion gene formed from a sperm motility kinase and a ribosomal S6 kinase. Three candidate distorter genes have also been identified as genes coding for dynein chains, and thus possibly involved in sperm flagellar function.

Gram-positive spore-forming entomopathogenic bacteria can utilize a large variety of protein toxins to help them invade, infect, and finally kill their hosts, through their action on the insect midgut. These toxins belong to a number of homology groups containing a diversity of protein structures and modes of action. In many cases, the toxins consist of unique folds or novel combinations of domains having known protein folds. Some of the toxins display a similar structure and mode of action to certain toxins of mammalian pathogens, suggesting a common evolutionary origin. Most of these toxins are produced in large amounts during sporulation and have the remarkable feature that they are localized in parasporal crystals. Localization of multiple toxin-encoding genes on plasmids together with mobilizable elements enables bacteria to shuffle their armory of toxins. Recombination between toxin genes and sequence divergence has resulted in a wide range of host specificities.

The vacuole/lysosome of the budding yeast Saccharomyces cerevisiae is actively divided between mother and daughter cells. Vacuole inheritance initiates early in the cell cycle and ends in G2, just prior to nuclear migration. The process begins with a portion of the vacuole extending into the emerging bud. This tubular-vesicular entity, the segregation structure, enables continued exchange of vacuole contents between mother and daughter vacuoles. Genetic, biochemical, and cytological analyses of vacuole inheritance have provided insight into the molecular basis of membrane movement, the spatial and temporal control of organelle transport, and the molecular basis of membrane fusion and fission.

Heparan sulfate (HS) binds numerous extracellular ligands, including cell-cell signaling molecules and their signal-transducing receptors. Ligand binding sites in HS have specific sulfation patterns; and several observations suggest that the HS sulfation pattern is the same for every HS chain that a cell synthesizes, regardless of the core protein to which it is attached. Nonetheless, virtually every Drosophila, zebrafish, Xenopus, and mouse that lacks a specific HS core protein has a mutant phenotype, even though other HS core proteins are expressed in the affected cells. Genetic manipulation of HS core protein genes is beginning to indicate that HS core proteins have functional specificities that are required during distinct stages of development.

Telomere molecular biology is far more complex than originally thought. Understanding biological systems is aided by study of evolutionary variants, and Drosophila telomeres are remarkable variants. Drosophila lack telomerase and the arrays of simple repeats generated by telomerase in almost all other organisms; instead, Drosophila telomeres are long tandem arrays of two non-LTR retrotransposons, HeT-A and TART. These are the first transposable elements found to have a bona fide role in cell structure, revealing an unexpected link between telomeres and what is generally considered to be parasitic DNA. In addition to providing insight into the cellular functions performed by telomeres, analysis of HeT-A and TART is providing insight into the evolution of chromosomes, retrotransposons, and retroviruses. Recent studies show that retrotransposon telomeres constitute a robust system for maintaining chromosome ends. These telomeres are now known to predate the separation of extant Drosophila species, allowing ample time for elements and hosts to coevolve interesting mechanisms.

Cyanobacteria such as Synechococcus elongatus PCC 7942 exhibit 24-h rhythms of gene expression that are controlled by an endogenous circadian clock that is mechanistically distinct from those described for diverse eukaryotes. Genetic and biochemical experiments over the past decade have identified key components of the circadian oscillator, input pathways that synchronize the clock with the daily environment, and output pathways that relay temporal information to downstream genes. The mechanism of the cyanobacterial circadian clock that is emerging is based principally on the assembly and disassembly of a large complex at whose heart are the proteins KaiA, KaiB, and KaiC. Signal transduction pathways that feed into and out of the clock employ protein domains that are similar to those in two-component regulatory systems of bacteria.