Genetic Model Organism Research Articles

Reproductive fitness and dietary choice behavior of the genetic model organism Caenorhabditis elegans under semi-natural conditions.

Laboratory breeding conditions of the model organism C. elegans do not correspond with the conditions in its natural soil habitat. To assess the consequences of the differences in environmental conditions, the effects of air composition, medium and bacterial food on reproductive fitness and/or dietary-choice behavior of C. elegans were investigated. The reproductive fitness of C. elegans was maximal under oxygen deficiency and not influenced by a high fractional share of carbon dioxide. In media approximating natural soil structure, reproductive fitness was much lower than in standard laboratory media. In seminatural media, the reproductive fitness of C. elegans was low with the standard laboratory food bacterium E. coli (γ-Proteobacteria), but significantly higher with C. arvensicola (Bacteroidetes) and B. tropica (β-Proteobacteria) as food. Dietary-choice experiments in semi-natural media revealed a low preference of C. elegans for E. coli but significantly higher preferences for C. arvensicola and B. tropica (among other bacteria). Dietary-choice experiments under quasi-natural conditions, which were feasible by fluorescence in situ hybridization (FISH) of bacteria, showed a high preference of C. elegans for Cytophaga-Flexibacter-Bacteroides, Firmicutes, and β-Proteobacteria, but a low preference for γ-Proteobacteria. The results show that data on C. elegans under standard laboratory conditions have to be carefully interpreted with respect to their biological significance.

Seasonal dynamics of a drosophilid (Diptera) assemblage and its potencial as bioindicator in open environments

Drosophila Fallen, 1823 (Diptera, Drosophilidae) is for long a well-established model organism for genetics and evolutionary research. The ecology of these flies, however, has only recently been better studied. Recent papers show that Drosophila assemblies can be used as bioindicators of forested environment degradation. In this work the bioindicator potential of drosophilids was evaluated in a naturally opened environment, a coastal strand-forest (restinga). Data from nine consecutive seasonal collections revealed strong temporal fluctuation pattern of the majority of Drosophila species groups. Drosophila willistoni group was more abundant at autumns, whereas D. cardini and D. tripunctata groups were, respectively, expressive at winters and springs, and D. repleta group at both seasons. The exotic species D. simulans Sturtevant, 1919 (from D. melanogaster group) and Zaprionus indianus Gupta, 1970 were most abundant at summers. Overall, the assemblage structure did not show the same characteristics of forested or urban environments, but was similar to the forests at winters and to cities at summers. This raises the question that this locality may already been under urbanization impact. Also, this can be interpreted as an easily invaded site for exotic species, what might lead to biotic homogenization and therefore can put in check the usage of drosophilid assemblages as bioindicators at open environments.

Rat traps: filling the toolbox for manipulating the rat genome

The laboratory rat is rapidly gaining momentum as a mammalian genetic model organism. Although traditional forward genetic approaches are well established, recent technological developments have enabled efficient gene targeting and mutant generation. Here we outline the current status, possibilities and application of these techniques in the rat.

Sordaria macrospora, a model organism to study fungal cellular development

During the development of multicellular eukaryotes, the processes of cellular growth and organogenesis are tightly coordinated. Since the 1940s, filamentous fungi have served as genetic model organisms to decipher basic mechanisms underlying eukaryotic cell differentiation. Here, we focus on Sordaria macrospora, a homothallic ascomycete and important model organism for developmental biology. During its sexual life cycle, S. macrospora forms three-dimensional fruiting bodies, a complex process involving the formation of different cell types. S. macrospora can be used for genetic, biochemical and cellular experimental approaches since diverse tools, including fluorescence microscopy, a marker recycling system and gene libraries, are available. Moreover, the genome of S. macrospora has been sequenced and allows functional genomics analyses. Over the past years, our group has generated and analysed a number of developmental mutants which has greatly enhanced our fundamental understanding about fungal morphogenesis. In addition, our recent research activities have established a link between developmental proteins and conserved signalling cascades, ultimately leading to a regulatory network controlling differentiation processes in a eukaryotic model organism. This review summarizes the results of our recent findings, thus advancing current knowledge of the general principles and paradigms underpinning eukaryotic cell differentiation and development.

Genetics and regulation of the major enzymes of alanine synthesis in Escherichia coli.

Genetic analysis of alanine synthesis in the model genetic organism Escherichia coli has implicated avtA, the still uncharacterized alaA and alaB genes, and probably other genes. We identified alaA as yfbQ. We then transferred mutations in several transaminase genes into a yfbQ mutant and isolated a mutant that required alanine for optimal growth. For cells grown with carbon sources other than pyruvate, the major alanine-synthesizing transaminases are AvtA, YfbQ (AlaA), and YfdZ (which we designate AlaC). Growth with pyruvate as the carbon source and multicopy suppression suggest that several other transaminases can contribute to alanine synthesis. Expression studies showed that alanine modestly repressed avtA and yfbQ but had no effect on yfdZ. The leucine-responsive regulatory protein (Lrp) mediated control by alanine. We purified YfbQ and YfdZ and showed that both are dimers with K(m)s for pyruvate within the intracellular range of pyruvate concentration.

Quantitative Proteomic Approaches and Their Application in the Study of Yeast Stress Responses

The increasing popularity of proteomics has created a need for quantitative analysis methods. As a result, many different techniques are now available for performing gel-based or gel-free quantitative proteomic analysis of biological samples. These techniques provide an insight into global protein expression from identification to quantification, taking into account the highly dynamic status of the proteome. With its relatively small and compact genome, yeast is widely used as a model organism for molecular genetics, cell biology, and proteomic studies. In this review, we will show how quantitative proteomics has been used to study the effect of different stresses on yeast.

Adult zebrafish as a model organism for behavioural genetics

Recent research has demonstrated the suitability of adult zebrafish to model some aspects of complex behaviour. Studies of reward behaviour, learning and memory, aggression, anxiety and sleep strongly suggest that conserved regulatory processes underlie behaviour in zebrafish and mammals. The isolation and molecular analysis of zebrafish behavioural mutants is now starting, allowing the identification of novel behavioural control genes. As a result of this, studies of adult zebrafish are now helping to uncover the genetic pathways and neural circuits that control vertebrate behaviour.

A natural genetic polymorphism affects retroactive interference in Drosophila melanogaster.

As environments change, animals update their internal representations of the external world. New information about the environment is learned and retained whereas outdated information is disregarded or forgotten. Retroactive interference (RI) occurs when the retrieval of previously learned information is less available owing to the acquisition of recently acquired information. Even though RI is thought to be a major cause of forgetting, its functional significance is still under debate. We find that natural allelic variants of the Drosophila melanogaster foraging gene known to affect rover and sitter behaviour differ in RI. More specifically, rovers who were previously shown to experience greater environmental heterogeneity while foraging display RI whereas sitters do not. Rover responses are biased towards more recent learning events. These results provide an ecological context to investigate the function of forgetting via RI and a suitable genetic model organism to address the evolutionary relevance of cognitive tasks.

Linking model systems to cancer therapeutics: the case of Mastermind

Genetics, and more recently genomics, reveal striking conservation in the fundamental signaling pathways that underlie normal and aberrant cell processes. Consequently, various genetic model organisms are now attracting the interest of biomedical scientists who are focused on therapeutic approaches to human disease. There are now several examples of studies in which Drosophila seems likely to facilitate advances in potential therapies, and a recent report has demonstrated the utility of the fly model for understanding and treating human disease. Basic developmental genetic information first obtained in Drosophila was used to design a therapeutic block to oncogenic Notch signaling that was associated with leukemia in mice. The story of Notch signaling in Drosophila demonstrates the potential for standard Drosophila molecular genetics in developing therapeutic strategies that are relevant to human disease.

Mushrooms: Morphological complexity in the fungi

The PNAS paper by Stajich et al. (1) describes a superbly assembled and annotated genome of one of the most morphologically complex fungi, the “inky cap” mushroom Coprinopsis cinerea, and illustrates the accomplishments that can be made when working with a genetic model organism and the challenges that lie ahead if we are to truly understand how genomes relate to the evolution of organism complexity (Fig. 1).

Two-photon calcium imaging from head-fixed Drosophila during optomotor walking behavior.

Drosophila melanogster is a model organism rich in genetic tools to manipulate and identify neural circuits involved in specific behaviors. Here we present a novel technique for two-photon calcium imaging in the central brain of head-fixed Drosophila walking on an air-supported ball. The ball’s motion is tracked at high resolution and can be treated as a proxy for the fly’s own movements. We used the genetically encoded calcium sensor, GCaMP3.0, to record from important elements of the motion-processing pathway, the horizontal-system (HS) lobula plate tangential cells (LPTCs) in the fly optic lobe. We presented motion stimuli to the tethered fly and found that calcium transients in HS-neurons correlated with robust optomotor behavior during walking. Our technique allows an entirely new set of questions to be addressed by monitoring behavior and physiology in identified neurons in a powerful genetic model organism with an extensive repertoire of walking behaviors.

I-SceI endonuclease: a new tool for DNA repair studies and genetic manipulations in streptomycetes

Actinomycetes are Gram-positive bacteria with a complex life cycle. They produce many pharmaceutically relevant secondary metabolites, including antibiotics and anticancer drugs. However, there is a limited number of biotechnological applications available as opposed to genetic model organisms like Bacillus subtilis or Escherichia coli. We report here a system for the functional expression of a synthetic gene encoding the I-SceI homing endonuclease in several streptomycetes. Using the synthetic sce(a) gene, we were able to create controlled genomic DNA double-strand breaks. A mutagenesis system, based on the homing endonuclease I-SceI, has been developed to construct targeted, non-polar, unmarked gene mutations in Streptomyces sp. Tü6071. In addition, we have shown that homologous recombination is a major pathway in streptomycetes to repair an I-SceI-generated DNA double-strand break. This novel I-SceI-based tool will be useful in fundamental studies on the repair mechanism of DNA double-strand breaks and for a variety of biotechnological applications.

Rapid identification of heterozygous mutations in Drosophila melanogaster using genomic capture sequencing

One of the key advantages of using Drosophila melanogaster as a genetic model organism is the ability to conduct saturation mutagenesis screens to identify genes and pathways underlying a given phenotype. Despite the large number of genetic tools developed to facilitate downstream cloning of mutations obtained from such screens, the current procedure remains labor intensive, time consuming, and costly. To address this issue, we designed an efficient strategy for rapid identification of heterozygous mutations in the fly genome by combining rough genetic mapping, targeted DNA capture, and second generation sequencing technology. We first tested this method on heterozygous flies carrying either a previously characterized dac(5) or sens(E2) mutation. Targeted amplification of genomic regions near these two loci was used to enrich DNA for sequencing, and both point mutations were successfully identified. When this method was applied to uncharacterized twr mutant flies, the underlying mutation was identified as a single-base mutation in the gene Spase18-21. This targeted-genome-sequencing method reduces time and effort required for mutation cloning by up to 80% compared with the current approach and lowers the cost to <$1000 for each mutant. Introduction of this and other sequencing-based methods for mutation cloning will enable broader usage of forward genetics screens and have significant impacts in the field of model organisms such as Drosophila.

Soma-germline interactions that influence germline proliferation in Caenorhabditis elegans.

Caenorhabditis elegans boasts a short lifecycle and high fecundity, two features that make it an attractive and powerful genetic model organism. Several recent studies indicate that germline proliferation, a prerequisite to optimal fecundity, is tightly controlled over the course of development. Cell proliferation control includes regulation of competence to proliferate, a poorly understood aspect of cell fate specification, as well as cell-cycle control. Furthermore, dynamic regulation of cell proliferation occurs in response to multiple external signals. The C. elegans germ line is proving a valuable model for linking genetic, developmental, systemic, and environmental control of cell proliferation. Here, we consider recent studies that contribute to our understanding of germ cell proliferation in C. elegans. We focus primarily on somatic control of germline proliferation, how it differs at different life stages, and how it can be altered in the context of the life cycle and changes in environmental status.

A Tyrosine-based Motif Localizes a Drosophila Vesicular Transporter to Synaptic Vesicles in Vivo

Vesicular neurotransmitter transporters must localize to synaptic vesicles (SVs) to allow regulated neurotransmitter release at the synapse. However, the signals required to localize vesicular proteins to SVs in vivo remain unclear. To address this question we have tested the effects of mutating proposed trafficking domains in Drosophila orthologs of the vesicular monoamine and glutamate transporters, DVMAT-A and DVGLUT. We show that a tyrosine-based motif (YXXY) is important both for DVMAT-A internalization from the cell surface in vitro, and localization to SVs in vivo. In contrast, DVGLUT deletion mutants that lack a putative C-terminal trafficking domain show more modest defects in both internalization in vitro and trafficking to SVs in vivo. Our data show for the first time that mutation of a specific trafficking motif can disrupt localization to SVs in vivo and suggest possible differences in the sorting of VMATs versus VGLUTs to SVs at the synapse.

Tips and tricks for preparing lampbrush chromosome spreads from Xenopus tropicalis oocytes

Due to their large size and fine organization, lampbrush chromosomes (LBCs) of amphibian oocytes have been for decades one of the favorite tools of biologists for the analysis of transcriptional and post-transcriptional processes at the cytological level. The emergence of the diploid Xenopus tropicalis amphibian as a model organism for vertebrate developmental genetics and the accumulation of sequence data made available by its recent genomic sequencing, strongly revive the interest of LBCs as a powerful tool to study genes expressed during oogenesis. We describe here a detailed protocol for preparing LBCs from X. tropicalis oocyte and give practical advice to encourage a large number of researchers to become familiar with these chromosomes.

C. elegans models of neuromuscular diseases expedite translational research

AbstractThe nematode Caenorhabditis elegans is a genetic model organism and the only animal with a complete nervous system wiring diagram. With only 302 neurons and 95 striated muscle cells, a rich array of mutants with defective locomotion and the facility for individual targeted gene knockdown by RNA interference, it lends itself to the exploration of gene function at nerve muscle junctions. With approximately 60% of human disease genes having a C. elegans homologue, there is growing interest in the deployment of lowcost, high-throughput, drug screens of nematode transgenic and mutant strains mimicking aspects of the pathology of devastating human neuromuscular disorders. Here we explore the contributions already made by C. elegans to our understanding of muscular dystrophies (Duchenne and Becker), spinal muscular atrophy, amyotrophic lateral sclerosis, Friedreich’s ataxia, inclusion body myositis and the prospects for contributions to other neuromuscular disorders. A bottleneck to low-cost, in vivo, large-scale chemical library screening for new candidate therapies has been rapid, automated, behavioural phenotyping. Recent progress in quantifying simple swimming (thrashing) movements is making such screening possible and is expediting the translation of drug candidates towards the clinic.

Physiology in Drosophila motion-sensitive neurons during walking and flight behavior

Event Abstract Back to Event Physiology in Drosophila motion-sensitive neurons during walking and flight behavior Johannes D. Seelig1*, Eugenia M. Chiappe1, Gus K. Lott1, Michael B. Reiser2 and Vivek Jayaraman1 1 HHMI, Janelia Farm Research Campus, United States 2 Janelia Farm Research Campus , United States Drosophila melanogaster is a genetic model organism with many experimental advantages, including the ability to genetically manipulate specific sub-populations of neurons. Selected neurons in the fruit fly central brain can also be targeted for in vivo electrophysiology and two-photon calcium imaging. This powerful combination of physiology and genetic tools-such as cell-type-specific labeling, activity sensing, light-activation and silencing-is increasingly being applied to questions in systems neuroscience [1]. Our goal is to understand circuit computations underlying sensory-motor transformation in the fly brain. This requires recording not just neural activity but also the fly's behavior. Towards this end, we will present two novel experimental setups that we have developed: (i) Two-photon calcium imaging while the fly is walking on an air-supported 'Buchner' ball [2, 3] in a virtual arena [4]. (ii) Two-photon calcium imaging when the tethered fly is flying in a virtual arena. In order to assess the quality of behavior in our imaging preparation, we focused on a well-known behavior, the optomotor response. This turning response to visual motion can be induced by presenting a large field vertical grating moving horizontally in front of the fly. We show that tethered walking and flying flies reliably perform optomotor behavior during two-photon imaging. To assess the quality of the calcium imaging, we investigated calcium responses in lobula plate tangential cells (LPTCs). This small group of neurons has been exhaustively studied in fixed preparations in larger flies, and their responses to large-field stimuli (such as those that induce the optomotor response) have been carefully characterized [5]. In blowflies, LPTCs respond to large field motion stimuli with strong calcium responses, making these cells ideal candidates to test the capabilities of the behaving Drosophila preparation. Recording from Horizontal System (HS) LPTCs, we find reliable calcium responses in behaving fruit flies using a new genetically encoded calcium indicator, GCaMP3.0 [6]. HS neurons show strong (~150±/F) and stable responses to motion in their preferred direction. These calcium responses correlate with the fly's own walking optomotor response in that direction. Do responses of HS neurons change depending on the behavioral state of the animal? We will present results that compare responses in walking flies to those in flying flies. In our poster, we will describe the details of setups that allow us to perform stable recordings from identified motion-sensitive interneurons in the visual system of walking and flying Drosophila. These recordings represent the first physiological recordings in behaving Drosophila and provide a platform for future explorations of decision-making and sensory-motor transformations in this powerful genetic model organism.

On the origin of amniotic stem cells: of mice and men

A common characteristic of mammals is the development of extraembryonic supporting tissues and organs that are required for embryonic implantation, survival and development in utero. The amnion is the innermost extraembryonic membrane that eventually surrounds the fetus of amniotes, and contains the amniotic fluid. Next to its function in in utero development, the amnion has been shown to have an important potential for clinical applications. It is mainly used as a dressing to stimulate healing in skin and ocular wounds. Moreover, cells derived from the amniotic membrane and amniotic fluid have been reported to possess stem cell features, like pluripotent differentiation ability. Little is known about the early development of this membrane in humans. The mouse is a powerful genetic model organism that can be used to address the dynamics and the developmental origin of amnion and amnion-derived stem cells. Here, we discuss some fundamental differences in amnion development in the disc-shaped primate embryo and in the cup-shaped mouse embryo. We emphasize the consequences that this may have on the derivation of amniotic "stem" cells. After revision of the different isolation procedures of amniotic (fluid) derived "stem" cells from rodents, we reveal striking differences in the sources used to derive these cells across studies. The profound differences in the development of the extraembryonic membranes and cavities between primates and rodents may result in comparing cell types of different developmental origins, eventually leading to missinterpretations.

Discovery and application of insertion-deletion (INDEL) polymorphisms for QTL mapping of early life-history traits in Atlantic salmon

BackgroundFor decades, linkage mapping has been one of the most powerful and widely used approaches for elucidating the genetic architecture of phenotypic traits of medical, agricultural and evolutionary importance. However, successful mapping of Mendelian and quantitative phenotypic traits depends critically on the availability of fast and preferably high-throughput genotyping platforms. Several array-based single nucleotide polymorphism (SNP) genotyping platforms have been developed for genetic model organisms during recent years but most of these methods become prohibitively expensive for screening large numbers of individuals. Therefore, inexpensive, simple and flexible genotyping solutions that enable rapid screening of intermediate numbers of loci (~75-300) in hundreds to thousands of individuals are still needed for QTL mapping applications in a broad range of organisms.ResultsHere we describe the discovery of and application of insertion-deletion (INDEL) polymorphisms for cost-efficient medium throughput genotyping that enables analysis of >75 loci in a single automated sequencer electrophoresis column with standard laboratory equipment. Genotyping of INDELs requires low start-up costs, includes few standard sample handling steps and is applicable to a broad range of species for which expressed sequence tag (EST) collections are available. As a proof of principle, we generated a partial INDEL linkage map in Atlantic salmon (Salmo salar) and rapidly identified a number of quantitative trait loci (QTLs) affecting early life-history traits that are expected to have important fitness consequences in the natural environment.ConclusionsThe INDEL genotyping enabled fast coarse-mapping of chromosomal regions containing QTL, thus providing an efficient means for characterization of genetic architecture in multiple crosses and large pedigrees. This enables not only the discovery of larger number of QTLs with relatively smaller phenotypic effect but also provides a cost-effective means for evaluation of the frequency of segregating QTLs in outbred populations which is important for further understanding how genetic variation underlying phenotypic traits is maintained in the wild.