RTEL1 maintains genomic stability by suppressing homologous recombination.

Homologous recombination (HR) is an important conserved process for DNA repair and ensures maintenance of genome integrity. Inappropriate HR causes gross chromosomal rearrangements and tumorigenesis in mammals. In yeast, the Srs2 helicase eliminates inappropriate recombination events, but the functional equivalent of Srs2 in higher eukaryotes has been elusive. Here, we identify C. elegans RTEL-1 as a functional analog of Srs2 and describe its vertebrate counterpart, RTEL1, which is required for genome stability and tumor avoidance. We find that rtel-1 mutant worms and RTEL1-depleted human cells share characteristic phenotypes with yeast srs2 mutants: lethality upon deletion of the sgs1/BLM homolog, hyperrecombination, and DNA damage sensitivity. In vitro, purified human RTEL1 antagonizes HR by promoting the disassembly of D loop recombination intermediates in a reaction dependent upon ATP hydrolysis. We propose that loss of HR control after deregulation of RTEL1 may be a critical event that drives genome instability and cancer.

REC-1 and HIM-5 distribute meiotic crossovers and function redundantly in meiotic double-strand break formation in Caenorhabditis elegans.

The Caenorhabditis elegans gene rec-1 was the first genetic locus identified in metazoa to affect the distribution of meiotic crossovers along the chromosome. We report that rec-1 encodes a distant paralog of HIM-5, which was discovered by whole-genome sequencing and confirmed by multiple genome-edited alleles. REC-1 is phosphorylated by cyclin-dependent kinase (CDK) in vitro, and mutation of the CDK consensus sites in REC-1 compromises meiotic crossover distribution in vivo. Unexpectedly, rec-1; him-5 double mutants are synthetic-lethal due to a defect in meiotic double-strand break formation. Thus, we uncovered an unexpected robustness to meiotic DSB formation and crossover positioning that is executed by HIM-5 and REC-1 and regulated by phosphorylation.

Rhabditophanes diutinus a parthenogenetic clade IV nematode with dauer larvae.

Comparative studies using non-parasitic model species such as Caenorhabditis elegans, have been very helpful in investigating the basic biology and evolution of parasitic nematodes. However, as phylogenetic distance increases, these comparisons become more difficult, particularly when outside of the nematode clade to which C. elegans belongs (V). One of the reasons C. elegans has nevertheless been used for these comparisons, is that closely related well characterized free-living species that can serve as models for parasites of interest are frequently not available. The Clade IV parasitic nematodes Strongyloides are of great research interest due to their life cycle and other unique biological features, as well as their medical and veterinary importance. Rhabditophanes, a closely related free-living genus, forms part of the Strongyloidoidea nematode superfamily. Rhabditophanes diutinus (= R. sp. KR3021) was included in the recent comparative genomic analysis of the Strongyloididae, providing some insight into the genomic nature of parasitism. However, very little is known about this species, limiting its usefulness as a research model. Here we provide a species description, name the species as R. diutinus and investigate its life cycle and subsequently gene expression in multiple life stages. We identified two previously unreported starvation induced life stages: dauer larvae and arrested J2 (J2A) larvae. The dauer larvae are morphologically similar to and are the same developmental stage as dauers in C. elegans and infective larvae in Strongyloides. As in C. elegans and Strongyloides, dauer formation is inhibited by treatment with dafachronic acid, indicating some genetic control mechanisms are conserved. Similarly, the expression patterns of putative dauer/infective larva control genes resemble each other, in particular between R. diutinus and Strongyloides spp. These findings illustrate and increase the usefulness of R. diutinus as a non-parasitic, easy to work with model species for the Strongyloididae for studying the evolution of parasitism as well as many aspects of the biology of Strongyloides spp, in particular the formation of infective larvae.

Genomic identification and functional analysis of essential genes in Caenorhabditis elegans.

BACKGROUND: Essential genes are required for an organism's viability and their functions can vary greatly, spreading across many pathways. Due to the importance of essential genes, large scale efforts have been undertaken to identify the complete set of essential genes and to understand their function. Studies of genome architecture and organization have found that genes are not randomly disturbed in the genome. RESULTS: Using combined genetic mapping, Illumina sequencing, and bioinformatics analyses, we successfully identified 44 essential genes with 130 lethal mutations in genomic regions of C. elegans of around 7.3 Mb from Chromosome I (left). Of the 44 essential genes, six of which were genes not characterized previously by mutant alleles, let-633/let-638 (B0261.1), let-128 (C53H9.2), let-511 (W09C3.4), let-162 (Y47G6A.18), let-510 (Y47G6A.19), and let-131 (Y71G12B.6). Examine essential genes with Hi-C data shows that essential genes tend to cluster within TAD units rather near TAD boundaries. We have also shown that essential genes in the left half of chromosome I in C. elegans function in enzyme and nucleic acid binding activities during fundamental processes, such as DNA replication, transcription, and translation. From protein-protein interaction networks, essential genes exhibit more protein connectivity than non-essential genes in the genome. Also, many of the essential genes show strong expression in embryos or early larvae stages, indicating that they are important to early development. CONCLUSIONS: Our results confirmed that this work provided a more comprehensive picture of the essential gene and their functional characterization. These genetic resources will offer important tools for further heath and disease research.

Spectrum of variations in dog-1/FANCJ and mdf-1/MAD1 defective Caenorhabditis elegans strains after long-term propagation.

BACKGROUND: Whole and partial chromosome losses or gains and structural chromosome changes are hallmarks of human tumors. Guanine-rich DNA, which has a potential to form a G-quadruplex (G4) structure, is particularly vulnerable to changes. In Caenorhabditis elegans, faithful transmission of G-rich DNA is ensured by the DOG-1/FANCJ deadbox helicase. RESULTS: To identify a spectrum of mutations, after long-term propagation, we combined whole genome sequencing (WGS) and oligonucleotide array Comparative Genomic Hybridization (oaCGH) analysis of a C. elegans strain that was propagated, in the absence of DOG-1 and MDF-1/MAD1, for a total of 470 generations, with samples taken for long term storage (by freezing) in generations 170 and 270. We compared the genomes of F170 and F470 strains and identified 94 substitutions, 17 InDels, 3 duplications, and 139 deletions larger than 20 bp. These homozygous variants were predicted to impact 101 protein-coding genes. Phenotypic analysis of this strain revealed remarkable fitness recovery indicating that mutations, which have accumulated in the strain, are not only tolerated but also cooperate to achieve long-term population survival in the absence of DOG-1 and MDF-1. Furthermore, deletions larger than 20 bp were the only variants that frequently occurred in G-rich DNA. We showed that 126 of the possible 954 predicted monoG/C tracts, larger than 14 bp, were deleted in unc-46 mdf-1 such-4; dog-1 F470 (JNC170). CONCLUSIONS: Here, we identified variants that accumulated in C. elegans' genome after long-term propagation in the absence of DOG-1 and MDF-1. We showed that DNA sequences, with G4-forming potential, are vulnerable to deletion-formation in this genetic background.

Synthetic lethality of cohesins with PARPs and replication fork mediators.

Synthetic lethality has been proposed as a way to leverage the genetic differences found in tumor cells to affect their selective killing. Cohesins, which tether sister chromatids together until anaphase onset, are mutated in a variety of tumor types. The elucidation of synthetic lethal interactions with cohesin mutants therefore identifies potential therapeutic targets. We used a cross-species approach to identify robust negative genetic interactions with cohesin mutants. Utilizing essential and non-essential mutant synthetic genetic arrays in Saccharomyces cerevisiae, we screened genome-wide for genetic interactions with hypomorphic mutations in cohesin genes. A somatic cell proliferation assay in Caenorhabditis elegans demonstrated that the majority of interactions were conserved. Analysis of the interactions found that cohesin mutants require the function of genes that mediate replication fork progression. Conservation of these interactions between replication fork mediators and cohesin in both yeast and C. elegans prompted us to test whether other replication fork mediators not found in the yeast were required for viability in cohesin mutants. PARP1 has roles in the DNA damage response but also in the restart of stalled replication forks. We found that a hypomorphic allele of the C. elegans SMC1 orthologue, him-1(e879), genetically interacted with mutations in the orthologues of PAR metabolism genes resulting in a reduced brood size and somatic cell defects. We then demonstrated that this interaction is conserved in human cells by showing that PARP inhibitors reduce the viability of cultured human cells depleted for cohesin components. This work demonstrates that large-scale genetic interaction screening in yeast can identify clinically relevant genetic interactions and suggests that PARP inhibitors, which are currently undergoing clinical trials as a treatment of homologous recombination-deficient cancers, may be effective in treating cancers that harbor cohesin mutations.

High-throughput capturing and characterization of mutations in essential genes of Caenorhabditis elegans.

BACKGROUND: Essential genes are critical for the development of all organisms and are associated with many human diseases. These genes have been a difficult category to study prior to the availability of balanced lethal strains. Despite the power of targeted mutagenesis, there are limitations in identifying mutations in essential genes. In this paper, we describe the identification of coding regions for essential genes mutated using forward genetic screens in Caenorhabditis elegans. The lethal mutations described here were isolated and maintained by a wild-type allele on a rescuing duplication. RESULTS: We applied whole genome sequencing to identify the causative molecular lesion resulting in lethality in existing C. elegans mutant strains. These strains are balanced and can be easily maintained for subsequent characterization. Our method can be effectively used to analyze mutations in a large number of essential genes. We describe here the identification of 64 essential genes in a region of chromosome I covered by the duplication sDp2. Of these, 42 are nonsense mutations, six are splice signal mutations, one deletion, and 15 are non-synonymous mutations. Many of the essential genes in this region function in cell cycle, transcriptional regulation, and RNA processing. CONCLUSIONS: The essential genes identified here are represented by mutant strains, many of which have more than one mutant allele. The genetic resource can be utilized to further our understanding of essential gene function and will be applicable to the study of C. elegans development, conserved cellular function, and ultimately lead to improved human health.

The ortholog of the human proto-oncogene ROS1 is required for epithelial development in C. elegans.

The orphan receptor ROS1 is a human proto-oncogene, mutations of which are found in an increasing number of cancers. Little is known about the role of ROS1, however in vertebrates it has been implicated in promoting differentiation programs in specialized epithelial tissues. In this study we show that the C. elegans ortholog of ROS1, the receptor tyrosine kinase ROL-3, has an essential role in orchestrating the morphogenesis and development of specialized epidermal tissues, highlighting a potentially conserved function in coordinating crosstalk between developing epithelial cells. We also provide evidence of a direct relationship between ROL-3, the mucin SRAP-1, and BCC-1, the homolog of mRNA regulating protein Bicaudal-C. This study answers a longstanding question as to the developmental function of ROL-3, identifies three new genes that are expressed and function in the developing epithelium of C. elegans, and introduces the nematode as a potentially powerful model system for investigating the increasingly important, yet poorly understood, human oncogene ROS1.

The atm-1 gene is required for genome stability in Caenorhabditis elegans.

The Ataxia-telangiectasia-mutated (ATM) gene in humans was identified as the basis of a rare autosomal disorder leading to cancer susceptibility and is now well known as an important signal transducer in response to DNA damage. An approach to understanding the conserved functions of this gene is provided by the model system, Caenorhabditis elegans. In this paper we describe the structure and loss of function phenotype of the ortholog atm-1. Using bioinformatic and molecular analysis we show that the atm-1 gene was previously misannotated. We find that the transcript is in fact a product of three gene predictions, Y48G1BL.2 (atm-1), K10E9.1, and F56C11.4 that together make up the complete coding region of ATM-1. We also characterize animals that are mutant for two available knockout alleles, gk186 and tm5027. As expected, atm-1 mutant animals are sensitive to ionizing radiation. In addition, however, atm-1 mutants also display phenotypes associated with genomic instability, including low brood size, reduced viability and sterility. We document several chromosomal fusions arising from atm-1 mutant animals. This is the first time a mutator phenotype has been described for atm-1 in C. elegans. Finally we demonstrate the use of a balancer system to screen for and capture atm-1-derived mutational events. Our study establishes C. elegans as a model for the study of ATM as a mutator potentially leading to the development of screens to identify therapeutic targets in humans.

Genomic sequence of a mutant strain of Caenorhabditis elegans with an altered recombination pattern.

BACKGROUND: The original sequencing and annotation of the Caenorhabditis elegans genome along with recent advances in sequencing technology provide an exceptional opportunity for the genomic analysis of wild-type and mutant strains. Using the Illumina Genome Analyzer, we sequenced the entire genome of Rec-1, a strain that alters the distribution of meiotic crossovers without changing the overall frequency. Rec-1 was derived from ethylmethane sulfonate (EMS)-treated strains, one of which had a high level of transposable element mobility. Sequencing of this strain provides an opportunity to examine the consequences on the genome of altering the distribution of meiotic recombination events. RESULTS: Using Illumina sequencing and MAQ software, 83% of the base pair sequence reads were aligned to the reference genome available at Wormbase, providing a 21-fold coverage of the genome. Using the software programs MAQ and Slider, we observed 1124 base pair differences between Rec-1 and the reference genome in Wormbase (WS190), and 441 between the mutagenized Rec-1 (BC313) and the wild-type N2 strain (VC2010). The most frequent base-substitution was G:C to A:T, 141 for the entire genome most of which were on chromosomes I or X, 55 and 31 respectively. With this data removed, no obvious pattern in the distribution of the base differences along the chromosomes was apparent. No major chromosomal rearrangements were observed, but additional insertions of transposable elements were detected. There are 11 extra copies of Tc1, and 8 of Tc2 in the Rec-1 genome, most likely the remains of past high-hopper activity in a progenitor strain. CONCLUSION: Our analysis of high-throughput sequencing was able to detect regions of direct repeat sequences, deletions, insertions of transposable elements, and base pair differences. A subset of sequence alterations affecting coding regions were confirmed by an independent approach using oligo array comparative genome hybridization. The major phenotype of the Rec-1 strain is an alteration in the preferred position of the meiotic recombination event with no other significant phenotypic consequences. In this study, we observed no evidence of a mutator effect at the nucleotide level attributable to the Rec-1 mutation.

Spectrum of mutational events in the absence of DOG-1/FANCJ in Caenorhabditis elegans.

The Caenorhabditis elegans ortholog of the Fanconi anemia pathway component J (FANCJ) is DOG-1, which is essential for genome stability. Previous studies have shown that disruption of the dog-1 gene generates small deletions of poly-C/poly-G tracts detectable by PCR and results in a mutator phenotype. In this paper, we describe the isolation and characterization of lethal mutations resulting from the loss of dog-1 function. The mutant strains were analyzed using a combination of techniques including genetic mapping, SNP mapping, and oaCGH (oligo array Comparative Genome Hybridization). Using the eT1 balancer system to recover lethal mutants, we isolated, in addition to small deletions, large chromosomal rearrangements, including duplications, translocations and deficiencies. The forward mutation frequency was 10-fold higher than the spontaneous frequency for eT1, and equivalent to that observed for low doses of standard mutagens. From a screen for suppressors of mdf-1/MAD1 lethality, we previously had isolated such-4(h2168), shown here to be a large tandem duplication. Thus, the range of mutational events caused by lack of DOG-1/FANCJ is much broader than previously described.

High-resolution array comparative genomic hybridization analysis reveals unanticipated complexity of genetic deficiencies on chromosome V in Caenorhabditis elegans.

Genomic rearrangements are widely used in Caenorhabditis elegans research but many remain incompletely characterized at the physical level. We have used oligo-array comparative genomic analysis to assess the physical structure of 20 deficiencies and a single duplication of chromosome V. We find that while deletions internal to the chromosome appear simple in structure, terminal deletions are complex, containing duplications in addition to the deletion. Additionally, we confirm that transposon-induced deficiencies contain breakpoints that initiate at Tc1 elements. Finally, 13 of these deficiencies are known to suppress recombination far beyond the extent of the deletion. These deficiencies fall into two classes: strong and weak suppressors of adjacent recombination. Analysis of the deleted regions in these deficiencies reveals no common physical sites to explain the observed differences in recombination suppression. However, we find a strong correlation between the size of the rearranged chromosome and the severity of recombination suppression. Rearranged chromosomes that have a minor effect on recombination fall within 2% of normal chromosome size. Our observations highlight the use of array-based approaches for the analysis of rearranged genomes, revealing previously unidentified deficiency characteristics and addressing biologically relevant questions.

The Caenorhabditis elegans vab-10 spectraplakin isoforms protect the epidermis against internal and external forces.

Morphogenesis of the Caenorhabditis elegans embryo is driven by actin microfilaments in the epidermis and by sarcomeres in body wall muscles. Both tissues are mechanically coupled, most likely through specialized attachment structures called fibrous organelles (FOs) that connect muscles to the cuticle across the epidermis. Here, we report the identification of new mutations in a gene known as vab-10, which lead to severe morphogenesis defects, and show that vab-10 corresponds to the C. elegans spectraplakin locus. Our analysis of vab-10 reveals novel insights into the role of this plakin subfamily. vab-10 generates isoforms related either to plectin (termed VAB-10A) or to microtubule actin cross-linking factor plakins (termed VAB-10B). Using specific antibodies and mutations, we show that VAB-10A and VAB-10B have distinct distributions and functions in the epidermis. Loss of VAB-10A impairs the integrity of FOs, leading to epidermal detachment from the cuticle and muscles, hence demonstrating that FOs are functionally and molecularly related to hemidesmosomes. We suggest that this isoform protects against forces external to the epidermis. In contrast, lack of VAB-10B leads to increased epidermal thickness during embryonic morphogenesis when epidermal cells change shape. We suggest that this isoform protects cells against tension that builds up within the epidermis.

Huntingtin-interacting protein 1 influences worm and mouse presynaptic function and protects Caenorhabditis elegans neurons against mutant polyglutamine toxicity.

Huntingtin-interacting protein 1 (HIP1) was identified through its interaction with htt (huntingtin), the Huntington's disease (HD) protein. HIP1 is an endocytic protein that influences transport and function of AMPA and NMDA receptors in the brain. However, little is known about its contribution to neuronal dysfunction in HD. We report that the Caenorhabditis elegans HIP1 homolog hipr-1 modulates presynaptic activity and the abundance of synaptobrevin, a protein involved in synaptic vesicle fusion. Presynaptic function was also altered in hippocampal brain slices of HIP1-/- mice demonstrating delayed recovery from synaptic depression and a reduction in paired-pulse facilitation, a form of presynaptic plasticity. Interestingly, neuronal dysfunction in transgenic nematodes expressing mutant N-terminal huntingtin was specifically enhanced by hipr-1 loss of function. A similar effect was observed with several other mutant proteins that are expressed at the synapse and involved in endocytosis, such as unc-11/AP180, unc-26/synaptojanin, and unc-57/endophilin. Thus, HIP1 is involved in presynaptic nerve terminal activity and modulation of mutant polyglutamine-induced neuronal dysfunction. Moreover, synaptic proteins involved in endocytosis may protect neurons against amino acid homopolymer expansion.

Synthetic lethal interactions identify phenotypic "interologs" of the spindle assembly checkpoint components.

Here, we report genetic interactions with mdf-1(gk2)/MAD1 in Caenorhabditis elegans. Nine are evolutionarily conserved or phenotypic "interologs" and two are novel enhancers, hcp-1 and bub-3. We show that HCP-1 and HCP-2, the two CENP-F-related proteins, recently implicated in the spindle assembly checkpoint (SAC) function, do not have identical functions, since hcp-1(RNAi), but not hcp-2(RNAi), enhances the lethality of the SAC mutants.

Suppressors of spindle checkpoint defect (such) mutants identify new mdf-1/MAD1 interactors in Caenorhabditis elegans.

The spindle assembly checkpoint (SAC) governs the timing of metaphase-to-anaphase transition and is essential for genome stability. The Caenorhabditis elegans mutant strain gk2 carries a deletion within the mdf-1/MAD1 gene that results in death of the homozygous strain after two or three generations. Here we describe 11 suppressors of the mdf-1(gk2) lethality, 10 identified in an ethyl methanesulfonate (EMS) mutagenesis screen and 1 isolated using the dog-1(gk10) (deletions of guanine-rich DNA) mutator strain. Using time-lapse imaging of early embryonic cells and germline mitotic division, we demonstrate that there are two classes of suppressors. Eight suppressors compensate for the loss of the checkpoint by delaying mitotic progression, which coincides with securin (IFY-1/Pds1) accumulation; three suppressors have normal IFY-1/Pds1 levels and normal anaphase onset. Furthermore, in the class of suppressors with delayed mitotic progression, we have identified four alleles of known suppressors emb-30/APC4 and fzy-1/CDC20, which are components of the anaphase-promoting complex/cyclosome (APC/C). In addition, we have identified another APC/C component capable of bypassing the checkpoint requirement that has not previously been described in C. elegans. The such-1/APC5-like mutation, h1960, significantly delays anaphase onset both in germline and in early embryonic cells.

Poly-G/poly-C tracts in the genomes of Caenorhabditis.

BACKGROUND: In the genome of Caenorhabditis elegans, homopolymeric poly-G/poly-C tracts (G/C tracts) exist at high frequency and are maintained by the activity of the DOG-1 protein. The frequency and distribution of G/C tracts in the genomes of C. elegans and the related nematode, C. briggsae were analyzed to investigate possible biological roles for G/C tracts. RESULTS: In C. elegans, G/C tracts are distributed along every chromosome in a non-random pattern. Most G/C tracts are within introns or are close to genes. Analysis of SAGE data showed that G/C tracts correlate with the levels of regional gene expression in C. elegans. G/C tracts are over-represented and dispersed across all chromosomes in another Caenorhabditis species, C. briggsae. However, the positions and distribution of G/C tracts in C. briggsae differ from those in C. elegans. Furthermore, the C. briggsae dog-1 ortholog CBG19723 can rescue the mutator phenotype of C. elegans dog-1 mutants. CONCLUSION: The abundance and genomic distribution of G/C tracts in C. elegans, the effect of G/C tracts on regional transcription levels, and the lack of positional conservation of G/C tracts in C. briggsae suggest a role for G/C tracts in chromatin structure but not in the transcriptional regulation of specific genes.

The Cdc20 homolog, FZY-1, and its interacting protein, IFY-1, are required for proper chromosome segregation in Caenorhabditis elegans.

Accurate chromosome segregation is achieved by a series of highly regulated processes that culminate in the metaphase-to-anaphase transition of the cell cycle. In the budding yeast Saccharomyces cerevisiae, the degradation of the securin protein Pds1 reverses the binding and inhibition of the separase protein Esp1. Esp1 cleaves Scc1. That cleavage promotes the dissociation of the cohesin complex from the chromosomes and leads the separation of sister chromatids. Proteolysis of Pds1 is regulated by the anaphase-promoting complex (APC), a large multi-subunit E3 ubiquitin ligase whose activity is regulated by Cdc20/Fizzy. We have previously shown that the Caenorhabditis elegans genes mdf-1/MAD1 and mdf-2/MAD2 encode key members of the spindle checkpoint. Loss of function of either gene leads to an accumulation of somatic and heritable defects and ultimately results in death. Here we show that a missense mutation in fzy-1/CDC20/Fizzy suppresses mdf-1 lethality. We identified a FZY-1-interacting protein, IFY-1, a novel destruction-box protein. IFY-1 accumulates in one-cell-arrested emb-30/APC4 embryos and interacts with SEP-1, a C. elegans separase, suggesting that IFY-1 functions as a C. elegans securin.

TOR deficiency in C. elegans causes developmental arrest and intestinal atrophy by inhibition of mRNA translation.

BACKGROUND: TOR is a phosphatidylinositol kinase (PIK)-related kinase that controls cell growth and proliferation in response to nutritional cues. We describe a C. elegans TOR homolog (CeTOR) and phenotypes associated with CeTOR deficiency. These phenotypes are compared with the response to starvation and the inactivation of a variety of putative TOR targets. RESULTS: Whether caused by mutation or RNA interference, TOR deficiency results in developmental arrest at mid-to-late L3, which is accompanied by marked gonadal degeneration and a pronounced intestinal cell phenotype. A population of refractile, autofluorescent intestinal vesicles, which take up the lysosomal dye Neutral Red, increases dramatically in size, while the number of normal intestinal vesicles and the intestinal cytoplasmic volume decrease progressively. This is accompanied by an increase in the gut lumen size and a compromise in the intestine's ability to digest and absorb nutrients. CeTOR-deficient larvae exhibit no significant dauer characteristics, but share some features with starved L3 larvae. Notably, however, starved larvae do not have severe intestinal atrophy. Inactivation of C. elegans p70S6K or TAP42 homologs does not reproduce CeTOR deficiency phenotypes, nor does inactivation of C. elegans TIP41, a putative negative regulator of CeTOR function, rescue CeTOR deficiency. In contrast, inactivating the C. elegans eIF-4G homolog and eIF-2 subunits results in developmental arrest accompanied by the appearance of large, refractile intestinal vesicles and severe intestinal atrophy resembling that of CeTOR deficiency. CONCLUSIONS: The developmental arrest and intestinal phenotypes of CeTOR deficiency are due to an inhibition of global mRNA translation. Thus, TOR is a major upstream regulator of overall mRNA translation in C. elegans, as in yeast.

Homologous recombination is required for genome stability in the absence of DOG-1 in Caenorhabditis elegans.

In C. elegans, DOG-1 prevents deletions that initiate in polyG/polyC tracts (G/C tracts), most likely by unwinding secondary structures that can form in G/C tracts during lagging-strand DNA synthesis. We have used the dog-1 mutant to assay the in vivo contribution of various repair genes to the maintenance of G/C tracts. Here we show that DOG-1 and the BLM ortholog, HIM-6, act synergistically during replication; simultaneous loss of function of both genes results in replicative stress and an increase in the formation of small deletions that initiate in G/C tracts. Similarly, we demonstrate that the C. elegans orthologs of the homologous recombination repair genes BARD1, RAD51, and XPF and the trans-lesion synthesis polymerases poleta and polkappa contribute to the prevention of deletions in dog-1 mutants. Finally, we provide evidence that the small deletions generated in the dog-1 background are not formed through homologous recombination, nucleotide excision repair, or nonhomologous end-joining mechanisms, but appear to result from a mutagenic repair mechanism acting at G/C tracts. Our data support the hypothesis that absence of DOG-1 leads to replication fork stalling that can be repaired by deletion-free or deletion-prone mechanisms.