Anchor cell signaling and vulval precursor cell positioning establish a reproducible spatial context during C. elegans vulval induction

How cells coordinate their spatial positioning through intercellular signaling events is poorly understood. Here we address this topic using Caenorhabditis elegans vulval patterning during which hypodermal vulval precursor cells (VPCs) adopt distinct cell fates determined by their relative positions to the gonadal anchor cell (AC). LIN-3/EGF signaling by the AC induces the central VPC, P6.p, to adopt a 1° vulval fate. Exact alignment of AC and VPCs is thus critical for correct fate patterning, yet, as we show here, the initial AC-VPC positioning is both highly variable and asymmetric among individuals, with AC and P6.p only becoming aligned at the early L3 stage. Cell ablations and mutant analysis indicate that VPCs, most prominently 1° cells, move towards the AC. We identify AC-released LIN-3/EGF as a major attractive signal, which therefore plays a dual role in vulval patterning (cell alignment and fate induction). Additionally, compromising Wnt pathway components also induces AC-VPC alignment errors, with loss of posterior Wnt signaling increasing stochastic vulval centering on P5.p. Our results illustrate how intercellular signaling reduces initial spatial variability in cell positioning to generate reproducible interactions across tissues

Carboxy-Terminal Truncation Activates glp-1 Protein to Specify Vulval Fates in Caenorhabditis elegans

The glp-1 and lin-12 genes encode homologous transmembrane proteins that may act as receptors for cell interactions during development. The glp-1 product is required for induction of germ-line proliferation and for embryogenesis. By contrast, lin-12 mediates somatic cell interactions, including those between the precursor cells that form the vulval hypodermis (VPCs). Here we analyse an unusual allele of glp-1, glp-1(q35), which displays a semidominant multivulva phenotype (Muv), as well as the typical recessive, loss-of-function Glp phenotypes (sterility and embryonic lethality). We find that the effects of glp-1(q35) on VPC development mimic those of dominant lin-12 mutations, even in the absence of lin-12 activity. The glp-1(q35) gene bears a nonsense mutation predicted to eliminate the 122 C-terminal amino acids, including a ProGluSerThr (PEST) sequence thought to destabilize proteins. We suggest that the carboxy terminus bears a negative regulatory domain which normally inactivates glp-1 in the VPCs. We propose that inappropriate glp-1(q35) activity can substitute for lin-12 to determine vulval fate, perhaps by driving the VPCs to proliferate

Predicting Phenotypic Diversity and the Underlying Quantitative Molecular Transitions

During development, signaling networks control the formation of multicellular patterns. To what extent quantitative fluctuations in these complex networks may affect multicellular phenotype remains unclear. Here, we describe a computational approach to predict and analyze the phenotypic diversity that is accessible to a developmental signaling network. Applying this framework to vulval development in C. elegans, we demonstrate that quantitative changes in the regulatory network can render ~500 multicellular phenotypes. This phenotypic capacity is an order-of-magnitude below the theoretical upper limit for this system but yet is large enough to demonstrate that the system is not restricted to a select few outcomes. Using metrics to gauge the robustness of these phenotypes to parameter perturbations, we identify a select subset of novel phenotypes that are the most promising for experimental validation. In addition, our model calculations provide a layout of these phenotypes in network parameter space. Analyzing this landscape of multicellular phenotypes yielded two significant insights. First, we show that experimentally well-established mutant phenotypes may be rendered using non-canonical network perturbations. Second, we show that the predicted multicellular patterns include not only those observed in C. elegans, but also those occurring exclusively in other species of the Caenorhabditis genus. This result demonstrates that quantitative diversification of a common regulatory network is indeed demonstrably sufficient to generate the phenotypic differences observed across three major species within the Caenorhabditis genus. Using our computational framework, we systematically identify the quantitative changes that may have occurred in the regulatory network during the evolution of these species. Our model predictions show that significant phenotypic diversity may be sampled through quantitative variations in the regulatory network without overhauling the core network architecture. Furthermore, by comparing the predicted landscape of phenotypes to multicellular patterns that have been experimentally observed across multiple species, we systematically trace the quantitative regulatory changes that may have occurred during the evolution of the Caenorhabditis genus

Microsporidia-nematode associations in methane seeps reveal basal fungal parasitism in the deep sea

The deep sea is Earth’s largest habitat but little is known about the nature of deep-sea parasitism. In contrast to a few characterized cases of bacterial and protistan parasites, the existence and biological significance of deep-sea parasitic fungi is yet to be understood. Here we report the discovery of a fungus-related parasitic microsporidium, Nematocenator marisprofundi n. gen. n. sp. that infects benthic nematodes at Pacific Ocean methane seeps on the Pacific Ocean floor. This infection is species-specific and has been temporally and spatially stable over two years of sampling, indicating an ecologically consistent host-parasite interaction. A high distribution of spores in the reproductive tracts of infected males and females and their absence from host nematodes’ intestines suggests a sexual transmission strategy in contrast to the fecal-oral transmission of most microsporidia. N. marisprofundi targets the host’s body wall muscles causing cell lysis, and in severe infection even muscle filament degradation. Phylogenetic analyses placed N. marisprofundi in a novel and basal clade not closely related to any described microsporidia clade, suggesting either that microsporidia-nematode parasitism occurred early in microsporidia evolution or that host specialization occurred late in an ancient deep-sea microsporidian lineage. Our findings reveal that methane seeps support complex ecosystems involving interkingdom interactions between bacteria, nematodes, and parasitic fungi and that microsporidia parasitism exists also in the deep sea biosphere

Visualization of C. elegans transgenic arrays by GFP

BACKGROUND: Targeting the green fluorescent protein (GFP) via the E. coli lac repressor (LacI) to a specific DNA sequence, the lac operator (lacO), allows visualization of chromosomes in yeast and mammalian cells. In principle this method of visualization could be used for genetic mosaic analysis, which requires cell-autonomous markers that can be scored easily and at single cell resolution. The C. elegans lin-3 gene encodes an epidermal growth factor family (EGF) growth factor. lin-3 is expressed in the gonadal anchor cell and acts through LET-23 (transmembrane protein tyrosine kinase and ortholog of EGF receptor) to signal the vulval precursor cells to generate vulval tissue. lin-3 is expressed in the vulval cells later, and recent evidence raises the possibility that lin-3 acts in the vulval cells as a relay signal during vulval induction. It is thus of interest to test the site of action of lin-3 by mosaic analysis. RESULTS: We visualized transgenes in living C. elegans by targeting the green fluorescent protein (GFP) via the E. coli lac repressor (LacI) to a specific 256 sequence repeat of the lac operator (lacO) incorporated into transgenes. We engineered animals to express a nuclear-localized GFP-LacI fusion protein. C. elegans cells having a lacO transgene result in nuclear-localized bright spots (i.e., GFP-LacI bound to lacO). Cells with diffuse nuclear fluorescence correspond to unbound nuclear localized GFP-LacI. We detected chromosomes in living animals by chromosomally integrating the array of the lacO repeat sequence and visualizing the integrated transgene with GFP-LacI. This detection system can be applied to determine polyploidy as well as investigating chromosome segregation. To assess the GFP-LacI•lacO system as a marker for mosaic analysis, we conducted genetic mosaic analysis of the epidermal growth factor lin-3, expressed in the anchor cell. We establish that lin-3 acts in the anchor cell to induce vulva development, demonstrating this method's utility in detecting the presence of a transgene. CONCLUSION: The GFP-LacI•lacO transgene detection system works in C. elegans for visualization of chromosomes and extrachromosomal transgenes. It can be used as a marker for genetic mosaic analysis. The lacO repeat sequence as an extrachromosomal array becomes a valuable technique allowing rapid, accurate determination of spontaneous loss of the array, thereby allowing high-resolution mosaic analysis. The lin-3 gene is required in the anchor cell to induce the epidermal vulval precursors cells to undergo vulval development

Bias and Evolution of the Mutationally Accessible Phenotypic Space in a Developmental System

Genetic and developmental architecture may bias the mutationally available phenotypic spectrum. Although such asymmetries in the introduction of variation may influence possible evolutionary trajectories, we lack quantitative characterization of biases in mutationally inducible phenotypic variation, their genotype-dependence, and their underlying molecular and developmental causes. Here we quantify the mutationally accessible phenotypic spectrum of the vulval developmental system using mutation accumulation (MA) lines derived from four wild isolates of the nematodes Caenorhabditis elegans and C. briggsae. The results confirm that on average, spontaneous mutations degrade developmental precision, with MA lines showing a low, yet consistently increased, proportion of developmental defects and variants. This result indicates strong purifying selection acting to maintain an invariant vulval phenotype. Both developmental system and genotype significantly bias the spectrum of mutationally inducible phenotypic variants. First, irrespective of genotype, there is a developmental bias, such that certain phenotypic variants are commonly induced by MA, while others are very rarely or never induced. Second, we found that both the degree and spectrum of mutationally accessible phenotypic variation are genotype-dependent. Overall, C. briggsae MA lines exhibited a two-fold higher decline in precision than the C. elegans MA lines. Moreover, the propensity to generate specific developmental variants depended on the genetic background. We show that such genotype-specific developmental biases are likely due to cryptic quantitative variation in activities of underlying molecular cascades. This analysis allowed us to identify the mutationally most sensitive elements of the vulval developmental system, which may indicate axes of potential evolutionary variation. Consistent with this scenario, we found that evolutionary trends in the vulval system concern the phenotypic characters that are most easily affected by mutation. This study provides an empirical assessment of developmental bias and the evolution of mutationally accessible phenotypes and supports the notion that such bias may influence the directions of evolutionary change

In Caenorhabditis elegans Nanoparticle-Bio-Interactions Become Transparent: Silica-Nanoparticles Induce Reproductive Senescence

While expectations and applications of nanotechnologies grow exponentially, little is known about interactions of engineered nanoparticles with multicellular organisms. Here we propose the transparent roundworm Caenorhabditis elegans as a simple but anatomically and biologically well defined animal model that allows for whole organism analyses of nanoparticle-bio-interactions. Microscopic techniques showed that fluorescently labelled nanoparticles are efficiently taken up by the worms during feeding, and translocate to primary organs such as epithelial cells of the intestine, as well as secondary organs belonging to the reproductive tract. The life span of nanoparticle-fed Caenorhabditis elegans remained unchanged, whereas a reduction of progeny production was observed in silica-nanoparticle exposed worms versus untreated controls. This reduction was accompanied by a significant increase of the ‘bag of worms’ phenotype that is characterized by failed egg-laying and usually occurs in aged wild type worms. Experimental exclusion of developmental defects suggests that silica-nanoparticles induce an age-related degeneration of reproductive organs, and thus set a research platform for both, detailed elucidation of molecular mechanisms and high throughput screening of different nanomaterials by analyses of progeny production

A Mechanistic Basis for the Coordinated Regulation of Pharyngeal Morphogenesis in Caenorhabditis elegans by LIN-35/Rb and UBC-18–ARI-1

Genetic redundancy, whereby two genes carry out seemingly overlapping functions, may in large part be attributable to the intricacy and robustness of genetic networks that control many developmental processes. We have previously described a complex set of genetic interactions underlying foregut development in the nematode Caenorhabditis elegans. Specifically, LIN-35/Rb, a tumor suppressor ortholog, in conjunction with UBC-18–ARI-1, a conserved E2/E3 complex, and PHA-1, a novel protein, coordinately regulates an early step of pharyngeal morphogenesis involving cellular re-orientation. Functional redundancy is indicated by the observation that lin-35; ubc-18 double mutants, as well as certain allelic combinations of pha-1 with either lin-35 or ubc-18, display defects in pharyngeal development, whereas single mutants do not. Using a combination of genetic and molecular analyses, we show that sup-35, a strong recessive suppressor of pha-1–associated lethality, also reverts the synthetic lethality of lin-35; ubc-18, lin-35; pha-1, and ubc-18 pha-1 double mutants. SUP-35, which contains C2H2-type Zn-finger domains as well as a conserved RMD-like motif, showed a dynamic pattern of subcellular localization during embryogenesis. We find that mutations in sup-35 specifically suppress hypomorphic alleles of pha-1 and that SUP-35, acting genetically upstream of SUP-36 and SUP-37, negatively regulates pha-1 transcription. We further demonstrate that LIN-35, a transcriptional repressor, and UBC-18–ARI-1, a complex involved in ubiquitin-mediated proteolysis, negatively regulate SUP-35 abundance through distinct mechanisms. We also show that HCF-1, a C. elegans homolog of host cell factor 1, functionally antagonizes LIN-35 in the regulation of sup-35. Our cumulative findings piece together the components of a novel regulatory network that includes LIN-35/Rb, which functions to control organ morphogenesis. Our results also shed light on general mechanisms that may underlie developmental genetic redundancies as well as principles that may govern complex disease traits

Current and Emerging Treatment Options for Castration-Resistant Prostate Cancer: A Focus on Immunotherapy

BACKGROUND: Castration-resistant prostate cancer is a disease with limited treatment options. However, the ongoing elucidation of the mechanisms underlying this disease continues to support the development of not only novel agents, but also innovative approaches. Among these therapies, immunotherapy has emerged as a promising strategy. DESIGN: This review article summarizes the most recent data from investigations of immunotherapies in castration-resistant prostate cancer (literature and congress searches current as of August 2011). RESULTS: Immunotherapeutic strategies such as passive immunization, vaccines, and particularly checkpoint blockade have demonstrated some efficacy as single agents. Elucidation of effective combinations of agents and drug regimens is ongoing but will require continued careful investigation, including the standardization of surrogate endpoints in clinical trials. CONCLUSIONS: It is hypothesized that the combination of immunotherapeutic agents with traditional and novel chemotherapeutics will potentiate the efficacy of the chemotherapeutics while maintaining manageable toxicity

To what extent does IQ 'explain' socio-economic variations in function?

<p>Abstract</p> <p>Background</p> <p>The aims of this study were to examine the extent to which higher intellectual abilities protect higher socio-economic groups from functional decline and to examine whether the contribution of intellectual abilities is independent of childhood deprivation and low birth weight and other socio-economic and developmental factors in early life.</p> <p>Methods</p> <p>The Maastricht Aging Study (MAAS) is a prospective cohort study based upon participants in a registration network of general practices in The Netherlands. Information was available on 1211 men and women, 24 – 81 years old, who were without cognitive impairment at baseline (1993 – 1995), who ever had a paid job, and who participated in the six-year follow-up. Main outcomes were longitudinal decline in important components of quality of life and successful aging, i.e., self-reported physical, affective, and cognitive functioning.</p> <p>Results</p> <p>Persons with a low occupational level at baseline showed more functional decline than persons with a high occupational level. Socio-economic and developmental factors from early life hardly contributed to the adult socio-economic differences in functional decline. Intellectual abilities, however, took into account more than one third of the association between adult socio-economic status and functional decline. The contribution of the intellectual abilities was independent of the early life factors.</p> <p>Conclusion</p> <p>Rather than developmental and socio-economic characteristics of early life, the findings substantiate the importance of intellectual abilities for functional decline and their contribution – as potential, but neglected confounders – to socio-economic differences in functioning, successful aging, and quality of life. The higher intellectual abilities in the higher socio-economic status groups may also underlie the higher prevalences of mastery, self-efficacy and efficient coping styles in these groups.</p