ObjectiveTo evaluate the diagnostic accuracy of high-risk human papillomavirus (hrHPV) assays on self samples and the efficacy of self sampling strategies to reach underscreened women.DesignUpdated meta-analysis.Data sourcesMedline (PubMed), Embase, and CENTRAL from 1 January 2013 to 15 April 2018 (accuracy review), and 1 January 2014 to 15 April 2018 (participation review).Review methodsAccuracy review: hrHPV assay on a vaginal self sample and a clinician sample; and verification of the presence of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) by colposcopy and biopsy in all enrolled women or in women with positive tests. Participation review: study population included women who were irregularly or never screened; women in the self sampling arm (intervention arm) were invited to collect a self sample for hrHPV testing; women in the control arm were invited or reminded to undergo a screening test on a clinician sample; participation in both arms was documented; and a population minimum of 400 women.Results56 accuracy studies and 25 participation trials were included. hrHPV assays based on polymerase chain reaction were as sensitive on self samples as on clinician samples to detect CIN2+ or CIN3+ (pooled ratio 0.99, 95% confidence interval 0.97 to 1.02). However, hrHPV assays based on signal amplification were less sensitive on self samples (pooled ratio 0.85, 95% confidence interval 0.80 to 0.89). The specificity to exclude CIN2+ was 2% or 4% lower on self samples than on clinician samples, for hrHPV assays based on polymerase chain reaction or signal amplification, respectively. Mailing self sample kits to the woman’s home address generated higher response rates to have a sample taken by a clinician than invitation or reminder letters (pooled relative participation in intention-to-treat-analysis of 2.33, 95% confidence interval 1.86 to 2.91). Opt-in strategies where women had to request a self sampling kit were generally not more effective than invitation letters (relative participation of 1.22, 95% confidence interval 0.93 to 1.61). Direct offer of self sampling devices to women in communities that were underscreened generated high participation rates (>75%). Substantial interstudy heterogeneity was noted (I2>95%).ConclusionsWhen used with hrHPV assays based on polymerase chain reaction, testing on self samples was similarly accurate as on clinician samples. Offering self sampling kits generally is more effective in reaching underscreened women than sending invitations. However, since response rates are highly variable among settings, pilots should be set up before regional or national roll out of self sampling strategies.

The precise machineries required for two aspects of eukaryotic DNA replication, Okazaki fragment processing (OFP) and telomere maintenance, are poorly understood. In this work, we present evidence that Saccharomyces cerevisiae Pif1 helicase plays a wider role in DNA replication than previously appreciated and that it likely functions in conjunction with Dna2 helicase/nuclease as a component of the OFP machinery. In addition, we show that Dna2, which is known to associate with telomeres in a cell-cycle-specific manner, may be a new component of the telomere replication apparatus. Specifically, we show that deletion of PIF1 suppresses the lethality of a DNA2-null mutant. The pif1⌬ dna2⌬ strain remains methylmethane sulfonate sensitive and temperature sensitive; however, these phenotypes can be suppressed by further deletion of a subunit of pol ␦, POL32. Deletion of PIF1 also suppresses the cold-sensitive lethality and hydroxyurea sensitivity of the pol32⌬ strain. Dna2 is thought to function by cleaving long flaps that arise during OFP due to excessive strand displacement by pol ␦ and/or by an as yet unidentified helicase. Thus, suppression of dna2⌬ can be rationalized if deletion of POL32 and/or PIF1 results in a reduction in long flaps that require Dna2 for processing. We further show that deletion of DNA2 suppresses the long-telomere phenotype and the high rate of formation of gross chromosomal rearrangements in pif1⌬ mutants, suggesting a role for Dna2 in telomere elongation in the absence of Pif1.Yeast Pif1 is the founding member of the Pif1 subfamily of superfamily 1 DNA helicases (3). While other organisms, such as Caenorhabditis elegans and Homo sapiens, have only one identified Pif1 family member, in yeast, there is a second, closely related, protein, Rrm3p (3). In yeast, neither of these helicases is essential and mutants lacking both are viable and repair proficient. Both yeast proteins have 5Ј-to-3Ј DNA helicase activity (21,30,31). The region of similarity between Pif1 and Rrm3 is limited to the seven helicase motifs, which exhibit 40% identity and 60% similarity (3). This may indicate that the two helicases have structurally similar DNA substrates. Nevertheless, the two helicases differ in their biological functions, and these differences are likely mediated not only by the helicase domain but also by the divergent N termini, which are not required for helicase activity (4). To date, it has been impossible to determine which helicase, Rrm3 or Pif1, is the functional homolog of the single ortholog in other eukaryotes.One difference between Rrm3 and Pif1 is in their function at the rRNA gene. In rrm3 mutants, there is an increase in replisome pausing at the Fob1 protein-bound replication fork barrier (RFB) in the rRNA gene (22). The hypothesis is that Rrm3 is required to remove proteins that block the fork at that point, since Rrm3 is required for promoting fork movement at over 1,400 loci in the yeast genome, in addition to the RFB. In contrast to Rrm3, Pif1 seems to be required for pausing at the rRNA...

Chromosome instability (CIN) is observed in most solid tumors and is linked to somatic mutations in genome integrity maintenance genes. The spectrum of mutations that cause CIN is only partly known and it is not possible to predict a priori all pathways whose disruption might lead to CIN. To address this issue, we generated a catalogue of CIN genes and pathways by screening ∼2,000 reduction-of-function alleles for 90% of essential genes in Saccharomyces cerevisiae. Integrating this with published CIN phenotypes for other yeast genes generated a systematic CIN gene dataset comprised of 692 genes. Enriched gene ontology terms defined cellular CIN pathways that, together with sequence orthologs, created a list of human CIN candidate genes, which we cross-referenced to published somatic mutation databases revealing hundreds of mutated CIN candidate genes. Characterization of some poorly characterized CIN genes revealed short telomeres in mutants of the ASTRA/TTT components TTI1 and ASA1. High-throughput phenotypic profiling links ASA1 to TTT (Tel2-Tti1-Tti2) complex function and to TORC1 signaling via Tor1p stability, consistent with the role of TTT in PI3-kinase related kinase biogenesis. The comprehensive CIN gene list presented here in principle comprises all conserved eukaryotic genome integrity pathways. Deriving human CIN candidate genes from the list allows direct cross-referencing with tumor mutational data and thus candidate mutations potentially driving CIN in tumors. Overall, the CIN gene spectrum reveals new chromosome biology and will help us to understand CIN phenotypes in human disease.

SignificanceCompetition among cooccurring bacteria can change the structure and function of a microbial community. However, little is known about the molecular mechanisms that impact such interactions in vivo. We used the association between bioluminescent bacteria and their squid host to study how environmentally transmitted bacteria compete for a limited number of host colonization sites. Our work suggests that Vibrio fischeri use a type VI secretion system, acting as a contact-dependent interbacterial “weapon,” to eliminate competing strains from cooccupying sites in the host. This work illuminates a mechanism by which strain-specific differences drive closely related bacteria to engage in lethal battles as they establish a beneficial symbiosis, revealing how genetic variation among potential colonizers directly impacts the spatial structure of the host-associated population.

In 1960, progressive sensorineural deafness (McKusick 304,700, DFN-1) was shown to be X-linked based on a description of a large Norwegian pedigree. More recently, it was shown that this original DFN-1 family represented a new type of recessive neurodegenerative syndrome characterized by postlingual progressive sensorineural deafness as the first presenting symptom in early childhood, followed by progressive dystonia, spasticity, dysphagia, mental deterioration, paranoia and cortical blindness. This new disorder, termed Mohr-Tranebjaerg syndrome (referred to here as DFN-1/MTS) was mapped to the Xq21.3-Xq22 region2. Using positional information from a patient with a 21-kb deletion in chromosome Xq22 and sensorineural deafness along with dystonia, we characterized a novel transcript lying within the deletion as a candidate for this complex syndrome. We now report small deletions in this candidate gene in the original DFN-1/MTS family, and in a family with deafness, dystonia and mental deficiency but not blindness. This gene, named DDP (deafness/ dystonia peptide), shows high levels of expression in fetal and adult brain. The DDP protein demonstrates striking similarity to a predicted Schizosaccharomyces pombe protein of no known function. Thus, is it likely that the DDP gene encodes an evolutionarily conserved novel polypeptide necessary for normal human neurological development.

Different types of gross chromosomal rearrangements (GCRs), including translocations, interstitial deletions, terminal deletions with de novo telomere additions, and chromosome fusions, are observed in many cancers. Multiple pathways, such as S-phase checkpoints, DNA replication, recombination, chromatin remodeling, and telomere maintenance that suppress GCRs have been identified. To experimentally expand our knowledge of other pathway(s) that suppress GCRs, we developed a generally applicable genome-wide screening method. In this screen, we identified 10 genes (ALO1, CDC50, CSM2, ELG1, ESC1, MMS4, RAD5, RAD18, TSA1, and UFO1) that encode proteins functioning in the suppression of GCRs. Moreover, the breakpoint junctions of GCRs from these GCR mutator mutants were determined with modified breakpoint-mapping methods. We also identified nine genes (AKR1, BFR1, HTZ1, IES6, NPL6, RPL13B, RPL27A, RPL35A, and SHU2) whose mutations generated growth defects with the pif1⌬ mutation. In addition, we found that some of these mutations changed the telomere size.

Ku86 plays a key role in nonhomologous end joining in organisms as evolutionarily disparate as bacteria and humans. In eukaryotic cells, Ku86 has also been implicated in the regulation of telomere length although the effect of Ku86 mutations varies considerably between species. Indeed, telomeres either shorten significantly, shorten slightly, remain unchanged, or lengthen significantly in budding yeast, fission yeast, chicken cells, or plants, respectively, that are null for Ku86 expression. Thus, it has been unclear which model system is most relevant for humans. We demonstrate here that the functional inactivation of even a single allele of Ku86 in human somatic cells results in profound telomere loss, which is accompanied by an increase in chromosomal fusions, translocations, and genomic instability. Together, these experiments demonstrate that Ku86, separate from its role in nonhomologous end joining, performs the additional function in human somatic cells of suppressing genomic instability through the regulation of telomere length.Most human tumors display some sort of chromosomal instability, ranging from minor DNA sequence changes to GCRs (gross chromosomal rearrangements) and aneuploidy (reviewed in references 53 and 68). These genomic alterations are often the causative event(s) in the transformation of a normal cell into a neoplastic cell. This occurs when the alteration results in the activation of proto-oncogenes or the inactivation of tumor suppressor genes and/or by the acquisition of a mutator phenotype (reviewed in reference 51). There are at least three proposed pathways by which chromosomal rearrangements may originate: (i) checkpoint defects (reviewed in reference 46), (ii) stalled replication fork collapse (reviewed in reference 15), and (iii) telomere dysfunction (reviewed in reference 54). Studies with yeast have demonstrated that a deficiency in any of these three pathways enhances chromosome loss and GCRs by up to 2 to 3 orders of magnitude (reviewed in reference 46). Similarly, in higher eukaryotes, mutations in genes regulating checkpoints and the repair of stalled replication forks result in a highly elevated frequency of GCRs (reviewed in references 38 and 78). Finally, in mice and humans, evidence is accumulating that the dysfunction of telomeres may be the driving force in the generation of genomic instability, which is strongly linked to cancer predisposition (reviewed in reference 54).Telomeres are the terminal structures of linear chromosomes. Telomeres appear to perform at least two functions: (i)

Symbiotic bacteria use diverse strategies to compete for host colonization sites. However, little is known about the environmental cues that modulate interbacterial competition as they transition between free-living and host-associated lifestyles. We used the mutualistic relationship between Eupyrmna scolopes squid and Vibrio fischeri bacteria to investigate how intraspecific competition is regulated as symbionts move from the seawater to a host-like environment. We recently reported that V. fischeri uses a type VI secretion system (T6SS) for intraspecific competition during host colonization. Here, we investigated how environmental viscosity impacts T6SS-mediated competition by using a liquid hydrogel medium that mimics the viscous host environment. Our data demonstrate that although the T6SS is functionally inactive when cells are grown under low-viscosity liquid conditions similar to those found in seawater, exposure to a host-like high-viscosity hydrogel enhances T6SS expression and sheath formation, activates T6SS-mediated killing in as little as 30 min, and promotes the coaggregation of competing genotypes. Finally, the use of mass spectrometry-based proteomics revealed insights into how cells may prepare for T6SS competition during this habitat transition. These findings, which establish the use of a new hydrogel culture condition for studying T6SS interactions, indicate that V. fischeri rapidly responds to the physical environment to activate the competitive mechanisms used during host colonization. IMPORTANCE Bacteria often engage in interference competition to gain access to an ecological niche, such as a host. However, little is known about how the physical environment experienced by free-living or host-associated bacteria influences such competition. We used the bioluminescent squid symbiont Vibrio fischeri to study how environmental viscosity impacts bacterial competition. Our results suggest that upon transition from a planktonic environment to a host-like environment, V. fischeri cells activate their type VI secretion system, a contact-dependent interbacterial nanoweapon, to eliminate natural competitors. This work shows that competitor cells form aggregates under host-like conditions, thereby facilitating the contact required for killing, and reveals how V. fischeri regulates a key competitive mechanism in response to the physical environment.