The Costs (and Benefits?) of Effortful Listening for Older Adults: Insights from Simultaneous Electrophysiology, Pupillometry, and Memory.

Although the impact of acoustic challenge on speech processing and memory increases as a person ages, older adults may engage in strategies that help them compensate for these demands. In the current preregistered study, older adults (n = 48) listened to sentences-presented in quiet or in noise-that were high constraint with either expected or unexpected endings or were low constraint with unexpected endings. Pupillometry and EEG were simultaneously recorded, and subsequent sentence recognition and word recall were measured. Like young adults in prior work, we found that noise led to increases in pupil size, delayed and reduced ERP responses, and decreased recall for unexpected words. However, in contrast to prior work in young adults where a larger pupillary response predicted a recovery of the N400 at the cost of poorer memory performance in noise, older adults did not show an associated recovery of the N400 despite decreased memory performance. Instead, we found that in quiet, increases in pupil size were associated with delays in N400 onset latencies and increased recognition memory performance. In conclusion, we found that transient variation in pupil-linked arousal predicted trade-offs between real-time lexical processing and memory that emerged at lower levels of task demand in aging. Moreover, with increased acoustic challenge, older adults still exhibited costs associated with transient increases in arousal without the corresponding benefits.

Small RNA in situ hybridization in Caenorhabditis elegans, combined with RNA-seq, identifies germline-enriched microRNAs.

Over four hundred different microRNAs (miRNAs) have been identified in the genome of the model organism the nematode Caenorhabditis elegans. As the germline is dedicated to the preservation of each species, and almost half of all the cells in an adult nematode are germline, it is likely that regulatory miRNAs are important for germline development and maintenance. In C. elegans the miR35 family has strong maternal effects, contributing to normal embryogenesis and to adult fecundity. To determine whether any particular miRNAs are greatly enriched in the C. elegans germline we used RNA-seq to compare the miRNA populations in several germline-defective strains of adult C. elegans worms, including glp-4(germline proliferation-4), glh-1(germline helicase-1) and dcr-1(dicer-1). Statistical analyses of RNA-seq comparisons identified 13 miRNAs that are germline-enriched, including seven members of the well-studied miR35 family that were reduced as much as 1000-fold in TaqMan qRT PCR miRNA assays. Along with the miR35s, six others: miR-56 (a member of the miR51 family),-70, -244, -260 , -788 and -4813, none of which previously considered as such, were also identified by RNA-seq as germline-enriched candidates. We went on to develop a successful miRNA in situ hybridization protocol for C. elegans, revealing miR35s specifically concentrate during oogenesis in the pachytene region of the gonad, and persist throughout early embryogenesis, while in adult animals neither let-7 nor miR-228 has a germline-bias.

Potential for natural and enhanced attenuation of sulphanilamide in a contaminated chalk aquifer.

Understanding antibiotic biodegradation is important to the appreciation of their fate and removal from the environment. In this research an Isotope Ratio Mass Spectrometry (IRMS) method was developed to evaluate the extent of biodegradation of the antibiotic, sulphanilamide, in contaminated groundwater. Results indicted an enrichment in δ13C of 8.44‰ from -26.56 (at the contaminant source) to -18.12‰ (300m downfield of the source). These results confirm reductions in sulphanilamide concentrations (from 650 to 10mg/L) across the contaminant plume to be attributable to biodegradation (56%) vs. other natural attenuation processes, such as dilution or dispersion (42%). To understand the controls on sulphanilamide degradation ex-situ microcosms assessed the influence of sulphanilamide concentration, redox conditions and an alternative carbon source. Results indicated, high levels of anaerobic capacity (~50% mineralisation) to degrade sulphanilamide under high (263mg/L), moderate (10mg/L) and low (0.02mg/L) substrate concentrations. The addition of electron acceptors; nitrate and sulphate, did not significantly enhance the capacity of the groundwater to anaerobically biodegrade sulphanilamide. Interestingly, where alternative carbon sources were present, the addition of nitrate and sulphate inhibited sulphanilamide biodegradation. These results suggest, under in-situ conditions, when a preferential carbon source was available for biodegradation, sulphanilamide could be acting as a nitrogen and/or sulphur source. These findings are important as they highlight sulphanilamide being used as a carbon and a putative nitrogen and sulphur source, under prevailing iron reducing conditions.

PAN-1, a P-granule component important for C. elegans fertility, has dual roles in the germline and soma.

In Caenorhabditis elegans, P granules are germline-specific, RNA-containing granules that segregate into the germline precursor cell during early embryogenesis. In this short report, PAN-1, which previously has been found by others in screens for genes causing larval molting defects, is identified here as a novel P-granule component and a binding partner of GLH-1 (Germline RNA Helicase-1), a constitutive, germline-specific, P-granule protein. The PAN-1 predicted protein contains multiple leucine-rich repeats (LRRs) and regions with similarities to F-box proteins. Antibodies raised against PAN-1 reveal it is present both in the soma and the germline. In the germline, PAN-1 uniquely localizes to P granules from the first larval stage onward and is unusual for a P-granule component in lacking recognizable RNA binding motifs. Homozygous pan-1(gk142) deletion worms arrest as larvae that are unable to molt and this phenotype is also seen with pan-1(RNAi) into wild type worms. pan-1(RNAi) into the somatic RNAi-defective strain rrf-1(pk1417) bypasses the larval arrest and allows an assessment of PAN-1 function in the germline. We find pan-1(RNAi) is variably effective in knocking down PAN-1 protein and results in adult progeny that display multiple germline defects. These phenocopies range from under-proliferation of the germline, as also seen with loss of GLH-1, to the induction of endomitotic replication in oocytes, both defects that result in sterility, to fertile animals with significantly reduced progeny numbers. Thus, while loss of PAN-1 in the soma inhibits molting, this report demonstrates that PAN-1 is also a P-granule component that is essential for fertility.

Building emotional intelligence: a strategy for emerging nurse leaders to reduce workplace bullying.

Bullying is one of the most concerning forms of aggression in health care organizations. Conceptualized as an emotion-based response, bullying is often triggered by today's workplace challenges. Unfortunately, workplace bullying is an escalating problem in nursing. Bullying contributes to unhealthy and toxic environments, which in turn contribute to ineffective patient care, increased stress, and decreased job satisfaction among health care providers. These equate to a poor workforce environment, which in turn increases hospital costs when nurses choose to leave. Nurse managers are in positions of power to recognize and address negative workplace behaviors, such as bullying. However, emerging leaders in particular may not be equipped with the tools to deal with bullying and consequently may choose to overlook it. Substantive evidence from other disciplines supports the contention that individuals with greater emotional intelligence are better equipped to recognize early signs of negative behavior, such as bullying. Therefore, fostering emotional intelligence in emerging nurse leaders may lead to less bullying and more positive workplace environments for nurses in the future.

C. elegans Dicer interacts with the P-granule component GLH-1 and both regulate germline RNPs.

P granules, ribonucleoprotein (RNP) complexes specific to the cytoplasmic side of the nuclear pores of Caenorhabditis elegans germ cells, are implicated in post-transcriptional control of maternally-transcribed mRNAs. Here we show a relationship in C. elegans of Dicer, the riboendonuclease processing enzyme of the RNA interference and microRNA pathways, with GLH-1, a germline-specific RNA helicase and a constitutive component of P granules. Based on results from GST-pull-downs and immunoprecipitations, GLH-1 binds DCR-1 and this binding does not require RNA. Both GLH-1 protein and glh-1 mRNA levels are reduced in the dcr-1(ok247) null mutant background; conversely, a reduction of DCR-1 protein is observed in the glh-1(gk100) deletion strain. Thus, in the C. elegans germline, DCR-1 and GLH-1 are interdependent. In addition, evidence indicates that DCR-1 protein levels, like those of GLH-1, are likely regulated by the Jun N-terminal kinase (JNK), KGB-1. In adult germ cells, DCR-1 is found in uniformly-distributed, small puncta both throughout the cytoplasm and the nucleus, on the inner side of nuclear pores, and associated with P granules. In arrested oocytes, GLH-1 and DCR-1 re-localize to cytoplasmic and cortically-distributed RNP granules and are necessary to recruit other components to these complexes. We predict that the GLH-1/DCR-1 complex may function in the transport, deposition, or regulation of maternally-transcribed mRNAs and their associated miRNAs.

RLE-1, an E3 ubiquitin ligase, regulates C. elegans aging by catalyzing DAF-16 polyubiquitination.

The forkhead transcription factor, DAF-16, a downstream target of the insulin/IGF-I signaling pathway in C. elegans, is indispensable both for lifespan regulation and stress resistance. The molecular mechanisms involved in regulating DAF-16 transcriptional activation remain undefined. Here, we have identified an E3 ubiquitin ligase, RLE-1 (regulation of longevity by E3), which regulates aging in C. elegans. Disruption of RLE-1 expression in C. elegans increases lifespan; this extension of lifespan is due to elevated DAF-16 protein but not to changes of daf-16 mRNA levels. We have also found that RLE-1 catalyzes DAF-16 ubiquitination, leading to degradation by the proteasome. Elimination of RLE-1 expression in C. elegans causes increased transcriptional activation and sustained nuclear localization of DAF-16. Overexpression of DAF-16 in rle-1 mutants increases worm lifespan, while disruption of DAF-16 expression in rle-1 mutants reverses their longevity. Thus, RLE-1 is an E3 ubiquitin ligase of DAF-16 that regulates C. elegans aging.

Increasing Diversity in Developmental Biology.

The demographic profile of the scientific and biomedical workforce in the United States does not reflect the population at large (https://ncses.nsf.gov/pubs/nsf21321/data-tables; www.census.gov), raising concerns that there will be too few trained researchers in the future, the scope of research interests will not be broad enough, gaps in equity and social justice will continue to increase, and the safeguards to the integrity of the scientific enterprise could be jeopardized. To diversify the pool of scientists, the Society for Developmental Biology (SDB) developed the Choose Development! Program-a two-summer immersion for undergraduate students belonging to underrepresented (UR) populations in STEM to join the research laboratory of an established SDB member. This research-intensive experience was augmented by a multi-tier mentoring plan for each student, society-wide recognition, professional development activities and networking at national meetings. The strengths of the Choose Development! Program were leveraged to expand inclusion and outreach at the Society's leadership level, the Board of Directors (BOD), which then led to significant changes that impacted the SDB community. The cumulative outcomes of the Choose Development! Program provides evidence that community-based, long-term advocacy, and mentoring of young UR scientists is successful in retaining UR students in scientific career paths and making a scientific society more inclusive.

Prevalence, nature and predictors of omitted medication doses in mental health hospitals: A multi-centre study.

OBJECTIVE: Limited evidence concerning the burden and predictors of omitted medication doses within mental health hospitals could severely limit improvement efforts in this specialist setting. This study aimed to determine the prevalence, nature and predictors of omitted medication doses affecting hospital inpatients in two English National Health Service (NHS) mental health trusts. METHODS: Over 6 data collection days trained pharmacy teams screened inpatient prescription charts for scheduled and omitted medication doses within 27 adult and elderly wards across 9 psychiatric hospitals. Data were collected for inpatients admitted up to two weeks prior to each data collection day. Omitted doses were classified as 'time critical' and 'preventable' based on established criteria. Omitted dose frequencies were presented with 95% confidence intervals (CI). Multilevel logistic regression analyses determined the predictors of omitted dose occurrence, with omission risks presented as adjusted odds ratios (OR) with 95% CI. RESULTS: 18,664 scheduled medication doses were screened for 444 inpatients and 2,717 omissions were identified, resulting in a rate of 14.6% (95% CI 14.1-15.1). The rate of 'time critical' omitted doses was 19.3% (95% CI 16.3-22.6%). 'Preventable' omitted doses comprised one third of all omissions (34.5%, 930/2694). Logistic regression analysis revealed that medicines affecting the central nervous system were 55% less likely to be omitted compared to all other medication classes (9.9% vs. 18.8%, OR 0.45 (0.40-0.52)) and that scheduled doses administered using non-oral routes were more likely to be omitted compared the oral route (inhaled OR 3.47 (2.64-4.57), topical 2.71 (2.11-3.46), 'other' 2.15 (1.19-3.90)). 'Preventable' dose omissions were more than twice as likely to occur for 'time critical' medications than non-time critical medications (50.4% vs. 33.8%, OR 2.24 (1.22-4.11)). CONCLUSIONS: Omitted medication doses occur commonly in mental health hospitals with 'preventable' omissions a key contributor to this burden. Important targets for remedial intervention have been identified.

The Dynamics of P Granule Liquid Droplets Are Regulated by the Caenorhabditis elegans Germline RNA Helicase GLH-1 via Its ATP Hydrolysis Cycle.

P granules are phase-separated liquid droplets that play important roles in the maintenance of germ cell fate in Caenorhabditis elegans Both the localization and formation of P granules are highly dynamic, but mechanisms that regulate such processes remain poorly understood. Here, we show evidence that the VASA-like germline RNA helicase GLH-1 couples distinct steps of its ATPase hydrolysis cycle to control the formation and disassembly of P granules. In addition, we found that the phenylalanine-glycine-glycine repeats in GLH-1 promote its localization at the perinucleus. Proteomic analyses of the GLH-1 complex with a GLH-1 mutation that interferes with P granule disassembly revealed transient interactions of GLH-1 with several Argonautes and RNA-binding proteins. Finally, we found that defects in recruiting the P granule component PRG-1 to perinuclear foci in the adult germline correlate with the fertility defects observed in various GLH-1 mutants. Together, our results highlight the versatile roles of an RNA helicase in controlling the formation of liquid droplets in space and time.

Genetic analysis of the Caenorhabditis elegans GLH family of P-granule proteins.

The Vasa DEAD-box helicases are widespread markers of germ cells across species, and in some organisms have been shown to be essential for germ-cell formation and development. In contrast to the single Vasa gene in most systems analyzed, Caenorhabditis elegans has four Vasa family members, the germline helicases GLH-1, GLH-2, GLH-3, and GLH-4. Our analysis of deletion alleles of each glh gene demonstrates that GLH-1 is the key member of the family: loss of GLH-1 function causes sterility that is mainly maternal effect, is manifested predominantly at elevated temperature, and is due to reduced germ-cell proliferation and impaired formation of both sperm and oocytes. The other GLHs are not essential. However, GLH-4 serves redundant roles with GLH-1: loss of both genes' function causes glh-1-like sterility at all temperatures. Molecular epistasis analysis demonstrates that GLH-1 and GLH-4 are required for proper association of the PGL family of proteins with P granules, suggesting a pathway of P-granule assembly in which the GLHs are upstream of the PGL proteins and the mRNA cap-binding protein IFE-1. While loss of some P-granule components causes worms to be defective in RNA interference, loss of GLH-1 and GLH-4 does not compromise RNAi. Thus, RNAi likely does not require intact P granules but instead relies on particular P-granule factors. We discuss the evolution of the Vasa/GLH genes and current views of their functions and the assembly and roles of germ granules among species.

What causes medication administration errors in a mental health hospital? A qualitative study with nursing staff.

OBJECTIVE: Medication administration errors (MAEs) are a common risk to patient safety in mental health hospitals, but an absence of in-depth studies to understand the underlying causes of these errors limits the development of effective remedial interventions. This study aimed to investigate the causes of MAEs affecting inpatients in a mental health National Health Service (NHS) hospital in the North West of England. METHODS: Registered and student mental health nurses working in inpatient psychiatric units were identified using a combination of direct advertisement and incident reports and invited to participate in semi-structured interviews utilising the critical incident technique. Interviews were designed to capture the participants' experiences of inpatient MAEs. All interviews were transcribed verbatim and subject to framework analysis to illuminate the underlying active failures, error/violation-provoking conditions and latent failures according to Reason's model of accident causation. RESULTS: A total of 20 participants described 26 MAEs (including 5 near misses) during the interviews. The majority of MAEs were skill-based slips and lapses (n = 16) or mistakes (n = 5), and were caused by a variety of interconnecting error/violation-provoking conditions relating to the patient, medicines used, medicines administration task, health care team, individual nurse and working environment. Some of these local conditions had origins in wider organisational latent failures. Recurrent and influential themes included inadequate staffing levels, unbalanced staff skill mix, interruptions/distractions, concerns with how the medicines administration task was approached and problems with communication. CONCLUSIONS: To our knowledge this is the first published in-depth qualitative study to investigate the underlying causes of specific MAEs in a mental health hospital. Our findings revealed that MAEs may arise due to multiple interacting error and violation provoking conditions and latent 'system' failures, which emphasises the complexity of this everyday task facing practitioners in clinical practice. Future research should focus on developing and testing interventions which address key local and wider organisational 'systems' failures to reduce error.

The Caenorhabditis elegans eukaryotic initiation factor 5A homologue, IFF-1, is required for germ cell proliferation, gametogenesis and localization of the P-granule component PGL-1.

Eukaryotic initiation factor 5A (eIF-5A) was originally isolated as a translation initiation factor. However, this function has since been reconsidered, with recent studies pointing to roles for eIF-5A in mRNA metabolism and trafficking [Microbiol. Mol. Biol. Rev. 66 (2002) 460; Eur. Mol. Biol. Org. J. 17 (1998) 2914]. The Caenorhabditis elegans genome contains two eIF-5A homologues, iff-1 and iff-2, whose functions in vivo were examined in this study. The iff-2 mutation causes somatic defects that include slow larval growth and disorganized somatic gonadal structures in hermaphrodites. iff-2 males show disorganized tail sensory rays and spicules. On the other hand, iff-1 mRNA is expressed in the gonad, and the lack of iff-1 activity causes sterility with an underproliferated germline resulting from impaired mitotic proliferation in both hermaphrodites and males. In spite of underproliferation, meiotic nuclei are observed, as revealed by presence of immunoreactivity to the anti-HIM-3 antibody; however, no gametogenesis occurs in the iff-1 gonads. These phenotypes are in part similar to the mutants affected in the components of P granules, which are the C. elegans counterparts of germ granules [Curr. Top Dev. Biol. 50 (2000) 155]. We found that localization of the P-granule component PGL-1 to P granules is disrupted in the iff-1 mutant. In summary, the two C. elegans homologues of eIF-5A act in different tissues: IFF-2 is required in the soma, and IFF-1 is required in the germline for germ cell proliferation, for gametogenesis after entry into meiosis, and for proper PGL-1 localization on P granules.

Assessment of Mycobacterium tuberculosis transmission in Oxfordshire, UK, 2007-12, with whole pathogen genome sequences: an observational study.

BACKGROUND: Patients born outside the UK have contributed to a 20% rise in the UK's tuberculosis incidence since 2000, but their effect on domestic transmission is not known. Here we use whole-genome sequencing to investigate the epidemiology of tuberculosis transmission in an unselected population over 6 years. METHODS: We identified all residents with Oxfordshire postcodes with a Mycobacterium tuberculosis culture or a clinical diagnosis of tuberculosis between Jan 1, 2007, and Dec 31, 2012, using local databases and checking against the national Enhanced Tuberculosis Surveillance database. We used Illumina technology to sequence all available M tuberculosis cultures from identified cases. Sequences were clustered by genetic relatedness and compared retrospectively with contact investigations. The first patient diagnosed in each cluster was defined as the index case, with links to subsequent cases assigned first by use of any epidemiological linkage, then by genetic distance, and then by timing of diagnosis. FINDINGS: Although we identified 384 patients with a diagnosis of tuberculosis, country of birth was known for 380 and we sequenced isolates from 247 of 269 cases with culture-confirmed disease. 39 cases were genomically linked within 13 clusters, implying 26 local transmission events. Only 11 of 26 possible transmissions had been previously identified through contact tracing. Of seven genomically confirmed household clusters, five contained additional genomic links to epidemiologically unidentified non-household members. 255 (67%) patients were born in a country with high tuberculosis incidence, conferring a local incidence of 109 cases per 100,000 population per year in Oxfordshire, compared with 3·5 cases per 100,000 per year for those born in low-incidence countries. However, patients born in the low-incidence countries, predominantly UK, were more likely to have pulmonary disease (adjusted odds ratio 1·8 [95% CI 1·2-2·9]; p=0·009), social risk factors (4·4 [2·0-9·4]; p<0·0001), and be part of a local transmission cluster (4·8 [1·6-14·8]; p=0·006). INTERPRETATION: Although inward migration has contributed to the overall tuberculosis incidence, our findings suggest that most patients born in high-incidence countries reactivate latent infection acquired abroad and are not involved in local onward transmission. Systematic screening of new entrants could further improve tuberculosis control, but it is important that health care remains accessible to all individuals, especially high-risk groups, if tuberculosis control is not to be jeopardised. FUNDING: UK Clinical Research Collaboration (Wellcome Trust, Medical Research Council, National Institute for Health Research [NIHR]), and NIHR Oxford Biomedical Research Centre.

C. elegans mitotic cyclins have distinct as well as overlapping functions in chromosome segregation.

Mitotic cyclins in association with the Cdk1 protein kinase regulate progression through mitosis in all eukaryotes. Here, we address to what extent mitotic cyclins in the nematode Caenorhabditis elegans provide overlapping functions or distinct biological activities. C. elegans expresses a single A-type cyclin (CYA-1), three typical B-type cyclins (CYB-1, CYB-2.1 and CYB-2.2), and one B3-subfamily member (CYB-3). While we observed clear redundancies between the cyb genes, cyb-1 and cyb-3 also contribute specific essential functions in meiosis and mitosis. CYB-1 and CYB-3 show similar temporal and spatial expression, both cyclins localize prominently to the nucleus, and both associate with CDK-1 and display histone H1 kinase activity in vitro. We demonstrate that inhibition of cyb-1 by RNAi interferes with chromosome congression and causes aneuploidy. In contrast, cyb-3(RNAi) embryos fail to initiate sister chromatid separation. Inhibition of both cyclins simultaneously results in a much earlier and more dramatic arrest. However, only the combination of cyb-1, cyb-3 and cyb-2.1/cyb-2.2 RNAi fully resembles cdk-1 inhibition. This combination of redundant and specific phenotypes supports that in vivo phosphorylation of certain Cdk targets can be achieved by multiple Cdk1/cyclin complexes, while phosphorylation of other targets requires a unique Cdk1/cyclin combination.

Increasing parent involvement in youth HIV prevention: a randomized Caribbean study.

This article presents preliminary findings of a randomized HIV prevention study in Trinidad and Tobago in the Caribbean. The study centers on a family HIV workshop aimed at strengthening parenting skills that are empirically linked to reducing adolescent HIV exposure and other sexual risks. These skills include parental monitoring; educating youth about HIV, sex, and other sexually transmitted infections (STI's); and discussing cultural and interpersonal pressures to have sex. Participants include 180 primary caregivers and their 12-14-year-old adolescents randomized to either the Trinidad and Tobago family HIV Workshop (N = 92) or a general workshop (N = 88). Intervention and control group participants completed pretest and posttest measures on parenting and HIV risk outcomes. Compared to controls, intervention parents reported improvements in HIV knowledge (d = .79); attitudes toward AIDS (d = .42); general communication with adolescents (d = .94); conversations with adolescents about sex (d = .95); conversations about sexual risks and values (d = .43); monitoring of adolescents (d = .34); conflicts with adolescents (d = .30); and intensity of daily parenting hassles (d = .35). Intervention and control parents did not differ in behavioral control, use of positive parenting techniques, and expansion of support networks. Implications for addressing rising HIV risks among young people in Trinidad and Tobago and the Caribbean are discussed.

GLH-1, the C. elegans P granule protein, is controlled by the JNK KGB-1 and by the COP9 subunit CSN-5.

The GLHs (germline RNA helicases) are constitutive components of the germline-specific P granules in the nematode Caenorhabditis elegans and are essential for fertility, yet how GLH proteins are regulated remains unknown. KGB-1 and CSN-5 are both GLH binding partners, previously identified by two-hybrid interactions. KGB-1 is a MAP kinase in the Jun N-terminal kinase (JNK) subfamily, whereas CSN-5 is a subunit of the COP9 signalosome. Intriguingly, although loss of either KGB-1 or CSN-5 results in sterility, their phenotypes are strikingly different. Whereas csn-5 RNA interference (RNAi) results in under-proliferated germlines, similar to glh-1/glh-4(RNAi), the kgb-1(um3) loss-of-function mutant exhibits germline over-proliferation. When kgb-1(um3) mutants are compared with wild-type C. elegans, GLH-1 protein levels are as much as 6-fold elevated and the organization of GLH-1 in P granules is grossly disrupted. A series of additional in vivo and in vitro tests indicates that KGB-1 and CSN-5 regulate GLH-1 levels, with GLH-1 targeted for proteosomal degradation by KGB-1 and stabilized by CSN-5. We propose the ;good cop: bad cop' team of CSN-5 and KGB-1 imposes a balance on GLH-1 levels, resulting in germline homeostasis. In addition, both KGB-1 and CSN-5 bind Vasa, a Drosophila germ granule component; therefore, similar regulatory mechanisms might be conserved from worms to flies.

The GLH proteins, Caenorhabditis elegans P granule components, associate with CSN-5 and KGB-1, proteins necessary for fertility, and with ZYX-1, a predicted cytoskeletal protein.

The GLH proteins belong to a family of four germline RNA helicases in Caenorhabditis elegans. These putative ATP-dependent enzymes localize to the P granules, which are nonmembranous complexes of protein and RNA exclusively found in the cytoplasm of all C. elegans germ cells and germ cell precursors. To determine what proteins the GLHs bind, C. elegans cDNA libraries were screened by the yeast two-hybrid method, using GLHs as bait. Three interacting proteins, CSN-5, KGB-1, and ZYX-1, were identified and further characterized. GST pull-down assays independently established that these proteins bind GLHs. CSN-5 is closely related to the subunit 5 protein of COP9 signalosomes, conserved multiprotein complexes of plants and animals. RNA interference (RNAi) with csn-5 results in sterile worms with small gonads and no oocytes, a defect essentially identical to that produced by RNAi with a combination of glh-1 and glh-4. KGB-1 is a putative JNK MAP kinase that GLHs bind. A kgb-1 deletion strain has a temperature-sensitive, sterile phenotype characterized by the absence of mature oocytes and the presence of trapped, immature oocytes that have undergone endoreplication. ZYX-1 is a LIM domain protein most like vertebrate Zyxin, a cytoskeletal adaptor protein. In C. elegans, while zyx-1 appears to be a single copy gene, neither RNAi depletion nor a zyx-1 deletion strain results in an obvious phenotype. These three conserved proteins are the first members in each of their families reported to associate with germline helicases. Similar to the loss of GLH-1 and GLH-4, loss of either CSN-5 or KGB-1 causes oogenesis to cease, but does not affect the initial assembly of P granules.