Molecular identification of a peroxidase gene controlling body size in the entomopathogenic nematode Steinernema hermaphroditum.

The entomopathogenic nematode Steinernema hermaphroditum was recently rediscovered and is being developed as a genetically tractable experimental system for the study of previously unexplored biology, including parasitism of its insect hosts and mutualism with its bacterial endosymbiont Xenorhabdus griffiniae. Through whole-genome re-sequencing and genetic mapping we have for the first time molecularly identified the gene responsible for a mutationally defined phenotypic locus in an entomopathogenic nematode. In the process we observed an unexpected mutational spectrum following ethyl methansulfonate mutagenesis in this species.

The entomopathogenic nematode Steinernema hermaphroditum is a self-fertilizing hermaphrodite and a genetically tractable system for the study of parasitic and mutualistic symbiosis.

Entomopathogenic nematodes (EPNs), including Heterorhabditis and Steinernema, are parasitic to insects and contain mutualistically symbiotic bacteria in their intestines (Photorhabdus and Xenorhabdus, respectively) and therefore offer opportunities to study both mutualistic and parasitic symbiosis. The establishment of genetic tools in EPNs has been impeded by limited genetic tractability, inconsistent growth in vitro, variable cryopreservation, and low mating efficiency. We obtained the recently described Steinernema hermaphroditum strain CS34 and optimized its in vitro growth, with a rapid generation time on a lawn of its native symbiotic bacteria Xenorhabdus griffiniae.

Comparative Epigenomics Reveals that RNA Polymerase II Pausing and Chromatin Domain Organization Control Nematode piRNA Biogenesis.

Piwi-interacting RNAs (piRNAs) are important for genome regulation across metazoans, but their biogenesis evolves rapidly. In Caenorhabditis elegans, piRNA loci are clustered within two 3-Mb regions on chromosome IV. Each piRNA locus possesses an upstream motif that recruits RNA polymerase II to produce an ∼28 nt primary transcript.

Improving the annotation of the Heterorhabditis bacteriophora genome.

Background: Genome assembly and annotation remain exacting tasks. As the tools available for these tasks improve, it is useful to return to data produced with earlier techniques to assess their credibility and correctness. The entomopathogenic nematode Heterorhabditis bacteriophora is widely used to control insect pests in horticulture.

A Toolkit of Engineered Recombinational Balancers in C. elegans.

Dejima and colleagues report using CRISPR/Cas9 to generate a new collection of greatly improved balancer chromosomes in the standard laboratory nematode Caenorhabditis elegans, using methods previously reported by the same laboratory, expanding the set of C. elegans balancers to cover nearly 90% of coding genes.

Transgene-free genome editing by germline injection of CRISPR/Cas RNA.

Genome modification by CRISPR/Cas offers its users the ability to target endogenous sites in the genome for cleavage and for engineering precise genomic changes using template-directed repair, all with unprecedented ease and flexibility of targeting. As such, CRISPR/Cas is just part of a set of recently developed and rapidly improving tools that offer great potential for researchers to functionally access the genomes of organisms that have not previously been extensively used in a laboratory setting. We describe in detail protocols for using CRISPR/Cas to target genes of experimental organisms, in a manner that does not require transformation to obtain transgenic lines and that should be readily applicable to a wide range of previously little-studied species.

Transgene-free genome editing in Caenorhabditis elegans using CRISPR-Cas.

CRISPR-Cas is an efficient method for genome editing in organisms from bacteria to human cells. We describe a transgene-free method for CRISPR-Cas-mediated cleavage in nematodes, enabling RNA-homology-targeted deletions that cause loss of gene function; analysis of whole-genome sequencing indicates that the nuclease activity is highly specific.

A systematic approach to multifactorial cardiovascular disease: causal analysis.

The combination of systems biology and large data sets offers new approaches to the study of cardiovascular diseases. These new approaches are especially important for the common cardiovascular diseases that have long been described as multifactorial. This promise is undermined by biologists' skepticism of the spider web-like network diagrams required to analyze these large data sets.

Functional specialization of sensory cilia by an RFX transcription factor isoform.

In animals, RFX transcription factors govern ciliogenesis by binding to an X-box motif in the promoters of ciliogenic genes. In Caenorhabditis elegans, the sole RFX transcription factor (TF) daf-19 null mutant lacks all sensory cilia, fails to express many ciliogenic genes, and is defective in many sensory behaviors, including male mating. The daf-19c isoform is expressed in all ciliated sensory neurons and is necessary and sufficient for activating X-box containing ciliogenesis genes.

The C. elegans protein CEH-30 protects male-specific neurons from apoptosis independently of the Bcl-2 homolog CED-9.

The developmental control of apoptosis is fundamental and important. We report that the Caenorhabditis elegans Bar homeodomain transcription factor CEH-30 is required for the sexually dimorphic survival of the male-specific CEM (cephalic male) sensory neurons; the homologous cells of hermaphrodites undergo programmed cell death. We propose that the cell-type-specific anti-apoptotic gene ceh-30 is transcriptionally repressed by the TRA-1 transcription factor, the terminal regulator of sexual identity in C.

A protocol describing pharynx counts and a review of other assays of apoptotic cell death in the nematode worm Caenorhabditis elegans.

Studies of the nematode worm Caenorhabditis elegans have provided important insights into the genetics of programmed cell death (PCD), and revealed molecular mechanisms conserved from nematodes to humans. The organism continues to offer opportunities to investigate the processes of apoptosis under very well-defined conditions and at single-cell resolution in living animals. Here, a survey of the common methods used to study the process of PCD in C.

The Caenorhabditis elegans F-box protein SEL-10 promotes female development and may target FEM-1 and FEM-3 for degradation by the proteasome.

The Caenorhabditis elegans F-box protein SEL-10 and its human homolog have been proposed to regulate LIN-12 Notch signaling by targeting for ubiquitin-mediated proteasomal degradation LIN-12 Notch proteins and SEL-12 PS1 presenilins, the latter of which have been implicated in Alzheimer's disease. We found that sel-10 is the same gene as egl-41, which previously had been defined by gain-of-function mutations that semidominantly cause masculinization of the hermaphrodite soma. Our results demonstrate that mutations causing loss-of-function of sel-10 also have masculinizing activity, indicating that sel-10 functions to promote female development.

New genes that interact with lin-35 Rb to negatively regulate the let-60 ras pathway in Caenorhabditis elegans.

Previous studies have shown that a synthetic multivulva phenotype results from mutations in genes that antagonize the ras-mediated intercellular signaling system responsible for vulval induction in Caenorhabditis elegans. Synthetic multivulva mutations define two classes of genes, A and B, and a mutation in a gene of each class is required to produce the multivulva phenotype. The ectopic vulval tissue in multivulva animals is generated by vulval precursor cells that in the wild type do not generate vulval tissue.