<Emphasis Type="Italic">odd-skipped</Emphasis> homologs function during gut development in<Emphasis Type="Italic"> C. elegans</Emphasis>

Genes in the odd-skipped (odd) family encode a discrete subset of C2H2 zinc finger proteins that are widely distributed among metazoan phyla. Although the initial member (odd) was identified as a Drosophila pair-rule gene, various homologs are expressed within each of the three germ layers in complex patterns that suggest roles in many pathways beyond segmentation. To further investigate the evolutionary history and extant functions of genes in this family, we have initiated a characterization of two homologs, odd-1 and odd-2, identified in the genome of the nematode, Caenorhabditis elegans. Sequence comparisons with homologs from insects (Drosophila and Anopheles) and mammals suggest that two paralogs were present within an ancestral metazoan; additional insect paralogs and both extant mammalian genes likely resulted from gene duplications that occurred after the split between the arthropods and chordates. Analyses of gene function using RNAi indicate that odd-1 and odd-2 play essential and distinct roles during gut development. Specific expression of both genes in the developing intestine and other cells in the vicinity of the gut was shown using GFP-reporters. These results indicate primary functions for both genes that are most like those of the Drosophila paralogs bowel and drumstick, and support a model in which gut specification represents the ancestral role for genes in this family.Edited by C. Desplan     

Strategies for automated analysis of <Emphasis Type="Italic">C. elegans</Emphasis> locomotion

Automated analysis of C. elegans behaviour is a rapidly developing field, offering the possibility of behaviour-based, high-throughput drug screens and systematic phenotyping. Standard methods for parameterizing worm shapes and movements are emerging, and progress has been made towards overcoming the difficulties introduced by interactions between worms, as well as worm coiling and omega turning. Current methods have facilitated the identification of subtle phenotypes and the characterisation of roles of neurones in forward locomotion and chemotaxis, as well as the quantitative characterisation of behaviour choice and circadian patterns of activity. Given the speed with which C. elegans has been deployed in genetic screens and chemical screens, it is to be hoped that wormtrackers may eventually provide similar rapidity in assaying behavioural phenotypes. However, considerable progress must be made before this can be accomplished. In the case of genome-wide RNAi screens, for example, the presence in the worm genome of some 19,000 genes means that even the minimal user intervention in an automatic phenotyping system will be very costly. Nonetheless, recent advances have shown that drug actions on large numbers of worms can be tracked, raising hopes that high-throughput behavioural screens may soon be available.     

Knockout of the folate transporter <Emphasis Type="Italic">folt-1</Emphasis> causes germline and somatic defects in <Emphasis Type="Italic">C. elegans</Emphasis>

Background  

The C. elegans gene folt-1 is an ortholog of the human reduced folate carrier gene. The FOLT-1 protein has been shown to transport folate and to be involved in uptake of exogenous folate by worms. A knockout mutation of the gene, folt-1(ok1460), was shown to cause sterility, and here we investigate the source of the sterility and the effect of the folt-1 knockout on somatic function.     

ANK repeat-domain of SHN-1 Is indispensable for <Emphasis Type="Italic">in vivo</Emphasis> SHN-1 function in <Emphasis Type="Italic">C. elegans</Emphasis>

Shank protein is one of the postsynaptic density (PSD) proteins which play a major role in proper localization of proteins at membranes. The shn-1, a homolog of Shank in Caenorhabditis elegans, is expressed in neurons, pharynx, intestine, vulva and sperm. We have previously reported a possible genetic interaction between Shank and IP₃ receptor by examining shn-1 RNAi in IP₃ receptor (itr-1) mutant background. In order to show the direct interaction of Shank and IP₃ receptor as well as to show the direct in vivo function of Shank, we have characterized two different mutant alleles of shn-1, which have different deletions in the different domains. shn-1 mutants were observed for Ca²⁺-related behavioral defects with itr-1 mutants. We found that only shn-1 mutant defective in ANK repeat-domain showed significant defects in defecation, pharyngeal pumping and fertility. In addition, we found that shn-1 regulates defecation, pharyngeal pumping and probably male fertility with itr-1. Thus, we suggest that Shank ANK repeat-domain along with PDZ may play a crucial role in regulating Ca²⁺-signaling with IP₃ receptor.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

Patterns of Gene Duplication in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> and <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

In this paper we present a new method for detecting block duplications in a genome. It is more stringent than previous ones in that it requires a more rigorous definition of paralogous genes and that it requires the paralogous proteins on the two blocks to be contiguous. In addition, it provides three criterion choices: (1) the same composition (i.e., having the same paralogues in the two windows), (2) the same composition and gene order, and (3) the same composition, gene order, and gene orientation. The method is completely automated, requiring no visual inspection as in previous methods. We applied it to analyze the complete genomes of S. cerevisiae and C. elegans. In yeast we detected fewer duplicated blocks than previously reported. In C. elegans, however, we detected more block duplications than previously reported, indicating that although our method has a more stringent definition of block duplication than previous ones, it may be more sensitive in detection because it considers every possible window rather than only fixed nonoverlapping windows. Our results show that block duplication is a common phenomenon in both organisms. The patterns of block duplication in the two species are, however, markedly different. The yeast shows much more extensive block duplication than the nematode, with some chromosomes having more than 40% of the duplications derived from block duplications. Moreover, in the yeast the majority of block duplications occurred between chromosomes, while in the nematode most block duplications occurred within chromosomes.     

Revision of the <Emphasis Type="Italic">Serrulati</Emphasis> species of <Emphasis Type="Italic">Penstemon</Emphasis> section <Emphasis Type="Italic">Saccanthera</Emphasis> subsection <Emphasis Type="Italic">Serrulati</Emphasis>

A revision of Penstemon sect. Saccanthera subsect. Serrulati includes a new species (P. salmonensis), a new variety (P. triphyllus var. infernalis), and the elevation of a subspecies to species (P. curtiflorus), bringing the total number of species to eight, which are keyed and described, complete with nomenclature and type citations.     

<Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> comes of age

A report of the European C. elegans 2008 meeting, Seville, Spain, 29 March-2 April 2008.     

The draft genome sequence of the nematode <Emphasis Type="Italic">Caenorhabditis briggsae</Emphasis>, a companion to <Emphasis Type="Italic">C. elegans</Emphasis>

The publication of the draft genome sequence of Caenorhabditis briggsae improves the annotation of the genome of its close relative Caenorhabditis elegans and will facilitate comparative genomics and the study of the evolutionary changes during development.     

The presynaptic machinery at the synapse of <Emphasis Type="Italic">C. elegans</Emphasis>

Synapses are specialized contact sites that mediate information flow between neurons and their targets. Important physical interactions across the synapse are mediated by synaptic adhesion molecules. These adhesions regulate formation of synapses during development and play a role during mature synaptic function. Importantly, genes regulating synaptogenesis and axon regeneration are conserved across the animal phyla. Genetic screens in the nematode Caenorhabditis elegans have identified a number of molecules required for synapse patterning and assembly. C. elegans is able to survive even with its neuronal function severely compromised. This is in comparison with Drosophila and mice where increased complexity makes them less tolerant to impaired function. Although this fact may reflect differences in the function of the homologous proteins in the synapses between these organisms, the most likely interpretation is that many of these components are equally important, but not absolutely essential, for synaptic transmission to support the relatively undemanding life style of laboratory maintained C. elegans. Here, we review research on the major group of synaptic proteins, involved in the presynaptic machinery in C. elegans, showing a strong conservation between higher organisms and highlight how C. elegans can be used as an informative tool for dissecting synaptic components, based on a simple nervous system organization.     

Usefulness of the Non-conventional <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> Model to Assess <Emphasis Type="Italic">Candida</Emphasis> Virulence

Invasive candidiasis is caused mainly by Candida albicans, but other Candida species have increasing etiologies. These species show different virulence and susceptibility levels to antifungal drugs. The aims of this study were to evaluate the usefulness of the non-conventional model Caenorhabditis elegans to assess the in vivo virulence of seven different Candida species and to compare the virulence in vivo with the in vitro production of proteinases and phospholipases, hemolytic activity and biofilm development capacity. One culture collection strain of each of seven Candida species (C. albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida metapsilosis, Candida orthopsilosis and Candida parapsilosis) was studied. A double mutant C. elegans AU37 strain (glp-4;sek-1) was infected with Candida by ingestion, and the analysis of nematode survival was performed in liquid medium every 24 h until 120 h. Candida establishes a persistent lethal infection in the C. elegans intestinal tract. C. albicans and C. krusei were the most pathogenic species, whereas C. dubliniensis infection showed the lowest mortality. C. albicans was the only species with phospholipase activity, was the greatest producer of aspartyl proteinase and had a higher hemolytic activity. C. albicans and C. krusei caused higher mortality than the rest of the Candida species studied in the C. elegans model of candidiasis.     

<Emphasis Type="Italic">Galleria</Emphasis><Emphasis Type="Italic">mellonella</Emphasis> are Resistant to <Emphasis Type="Italic">Pneumocystis</Emphasis><Emphasis Type="Italic">murina</Emphasis> Infection

Studying Pneumocystis has proven to be a challenge from the perspective of propagating a significant amount of the pathogen in a facile manner. The study of several fungal pathogens has been aided by the use of invertebrate model hosts. Our efforts to infect the invertebrate larvae Galleria mellonella with Pneumocystis proved futile since P. murina neither caused disease nor was able to proliferate within G. mellonella. It did, however, show that the pathogen could be rapidly cleared from the host.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

Diverse roles of actin in <Emphasis Type="Italic">C. elegans</Emphasis>early embryogenesis

Background  

The actin cytoskeleton plays critical roles in early development in Caenorhabditis elegans. To further understand the complex roles of actin in early embryogenesis we use RNAi and in vivo imaging of filamentous actin (F-actin) dynamics.     

Identifying novel genes in <Emphasis Type="Italic">C. elegans</Emphasis> using SAGE tags

Background  

Despite extensive efforts devoted to predicting protein-coding genes in genome sequences, many bona fide genes have not been found and many existing gene models are not accurate in all sequenced eukaryote genomes. This situation is partly explained by the fact that gene prediction programs have been developed based on our incomplete understanding of gene feature information such as splicing and promoter characteristics. Additionally, full-length cDNAs of many genes and their isoforms are hard to obtain due to their low level or rare expression. In order to obtain full-length sequences of all protein-coding genes, alternative approaches are required.