The Thioredoxin TRX-1 Modulates the Function of the Insulin-Like Neuropeptide DAF-28 during Dauer Formation in Caenorhabditis elegans

Thioredoxins comprise a conserved family of redox regulators involved in many biological processes, including stress resistance and aging. We report that the C. elegans thioredoxin TRX-1 acts in ASJ head sensory neurons as a novel modulator of the insulin-like neuropeptide DAF-28 during dauer formation. We show that increased formation of stress-resistant, long-lived dauer larvae in mutants for the gene encoding the insulin-like neuropeptide DAF-28 requires TRX-1 acting in ASJ neurons, upstream of the insulin-like receptor DAF-2. Genetic rescue experiments demonstrate that redox-independent functions of TRX-1 specifically in ASJ neurons are needed for the dauer formation constitutive (Daf-c) phenotype of daf-28 mutants. GFP reporters of trx-1 and daf-28 show opposing expression patterns in dauers (i.e. trx-1 is up-regulated and daf-28 is down-regulated), an effect that is not observed in growing L2/L3 larvae. In addition, functional TRX-1 is required for the down-regulation of a GFP reporter of daf-28 during dauer formation, a process that is likely subject to DAF-28-mediated feedback regulation. Our findings demonstrate that TRX-1 modulates DAF-28 signaling by contributing to the down-regulation of daf-28 expression during dauer formation. We propose that TRX-1 acts as a fluctuating neuronal signaling modulator within ASJ neurons to monitor the adjustment of neuropeptide expression, including insulin-like proteins, during dauer formation in response to adverse environmental conditions.

Analysis of Centrosome duplication in "Caenorhabditis elegans" : the role of "sas" genes

Abstract: The centrosome is the major microtubule organizing center (MTOC) of most animal cells. As such, it is essential for a number of processes, including polarized secretion or bipolar spindle assembly. Hence, centrosome number needs to be controlled precisely in coordination with DNA replication. Cells early in the cell cycle contain one centrosome that duplicates during S-phase to give rise to two centrosomes that organize a bipolar spindle during mitosis. A failure in this process is likely to engage the spindle assembly checkpoint and threaten genome stability. Despite its importance for normal and uncontrolled proliferation the mechanisms underlying centrosome duplication are still unclear. The Caenorhabditis elegans embryo is well suited to study the mechanisms of centrosome duplication. It allows for the analysis of cellular processes with high temporal and spatial resolution. Gene identification and inactivation techniques are very powerful and a wide set of mutant and transgenic strains facilitates analysis. My thesis project consisted of characterizing three sas-genes: sas-4, sas-5 and sas-¬6. Embryos lacking these genes fail to form a bipolar spindle, hence their name (spindle assembly). I established that sas-4(RNAi) and sas-6(RNAi) embryos do not form daughter centrioles and thus do not duplicate their centrosomes. Furthermore, I showed that both proteins localize to the cytoplasm and are strikingly enriched at centrioles throughout the cell cycle. By performing fluorescent recovery after photobleaching (FRAP) experiments and differentially labeling centrioles, I established that both proteins are recruited to centrioles once per cell cycle when daughter centrioles form. In contrast, SAS-5, PLK-1 and SPD-2 shuttle permanently between the cytoplasm and centrioles. By showing that SAS-5 and SAS-6 interact in vivo, I established a functional relationship between the proteins. Testing the putative human homologue of SAS-6 (HsSAS-6) and a distant relative of SAS-4 (CPAP), I was able to show that these proteins are required for centrosome duplication in human cells. In addition I found that overexpression of GFP¬HsSAS-6 leads to formation of extra centrosomes. In conclusion, we identified and gained important insights into proteins required for centrosome duplication in C. elegans and in human cells. Thus, our work contributes to further elucidate an important step of cell division in normal and malignant tissues. Eventually, this may allow for the development of novel diagnostic or therapeutic reagents to treat cancer patients. Résumé: Le centrosome est le principal centre organisateur des microtubules dans les cellules animales. De ce fait, il est essentiel pour un certain nombre de processus, comme l'adressage polarisé ou la mise en place d'un fuseau bipolaire. Le nombre de centrosome doit être contrôlé de façon précise et en coordination avec la réplication de l'ADN. Au début du cycle cellulaire, les cellules n'ont qu'un seul centrosome qui se duplique au cours de la phase S pour donner naissance à deux centrosomes qui forment le fuseau bipolaire pendant la mitose. Des défauts dans ce processus déclencheront probablement le "checkpoint" d'assemblage du fuseau et menaceront la stabilité du génome. Malgré leurs importances pour la prolifération normale ou incontrôlée des cellules, les mécanismes gouvernant la duplication des centrosomes restent obscures. L'embryon de Caenorhabditis elegans est bien adapté pour étudier les mécanismes de duplication des centrosomes. Il permet l'analyse des processus cellulaires avec une haute résolution spatiale et temporelle. L'identification des gènes et les techniques d'inactivation sont très puissantes et de larges collections de mutants et de lignées transgéniques facilitent les analyses. Mon projet de thèse a consisté à caractérisé trois gènes: sas-4, sas-5 et sas-6. Les embryons ne possédant pas ces gènes ne forment pas de fuseaux bipolaires, d'où leur nom (spindle assembly). J'ai établi que les embryons sas-4(RNAi) et sas-6(RNAi) ne forment pas de centrioles fils, et donc ne dupliquent pas leur centrosome. De plus, j'ai montré que les deux protéines sont localisées dans le cytoplasme et sont étonnamment enrichies aux centrioles tout le long du cycle cellulaire. En réalisant des expériences de FRAP (fluorscence recovery after photobleaching) et en marquant différentiellement les centrioles, j'ai établi que ces deux protéines sont recrutées une fois par cycle cellulaire aux centrioles, au moment de la duplication. Au contraire, SAS-5, PLK-1 et SPD-2 oscillent en permanence entre le cytoplasme et les centrioles. En montrant que SAS-5 et SAS-6 interagissent in vivo, j'ai établi une relation fonctionnelle entre les deux protéines. En testant les homologues humains putatifs de SAS-6 (HsSAS-6) et de SAS-4 (CPAP), j'ai été capable de montrer que ces protéines étaient aussi requises pour la duplication des centrosomes dans les cellules humaines. De plus, j'ai montré que la surexpression de GFP-HsSAS-6 entrainait la formation de centrosomes surnuméraires. En conclusion, nous avons identifié et progressé dans la compréhension de protéines requises pour la duplication des centrosomes chez C. elegans et dans les cellules humaines. Ainsi, notre travail contribue à mieux élucider une étape importante du la division cellulaire dans les cellules normales et malignes. A terme, ceci devrait aider au développement de nouveaux diagnostics ou de traitements thérapeuthiques pour soigner les malades du cancer.

zyg-11and cul-2 promote the metaphase to anaphase transition and M phase exit of meiosis II and prevent polarity establishment in C.elegans

Résumé Les mécanismes qui coordonnent la progression du cycle cellulaire lors de la méiose avec les événements du développement embryonnaire précoce, y compris la formation des axes de polarité embryonnaire, sont peu compris. Dans le zygote du vers Caenorhabditis elegans, les premiers signes de polarité Antéro-Postérieur (A-P) embryonnaire apparaissent après que la méiose soit terminée. La nature des protéines et des mécanismes moléculaires qui cassent la symétrie du zygote n'est pas connue. Nous démontrons que zyg-11 et cul-2 promeuvent la transition métaphase - anaphase et la sortie de la phase M lors de la seconde division méiotique. Nos résultats indiquent que ZYG-11 agit comme unité recrutant le substrat d'une ligase E3 comprennant CUL-2. Nos résultats montrent aussi que le délai de sortie de la phase M dépend de l'accumulation de la Cyclin B, CYB-3. Nous démontrons que dans des embryons zyg-11(RNAi) ou cul-2(RNAi), une polarité inversée est établie lors du délai de méiosis II. Enfin nous montrons que les défauts de cycle cellulaire et ceux de polarité peuvent être séparés. De plus, nous faisons apparaitre que l'établissement d'une polarité inversée pendant le délai de méiose II des embryons zyg-11(RNAi), comme l'établissement de la A-P polarité des embryons sauvage ne semblent pas requérir les microtubules. Nous montrons également les premiers résultats d'un crible deux hybrides ainsi qu'un crible génomique qui vise à identifier des gènes dont l'inactivation augmente ou supprime les défauts de mutants pour le gène zyg-11, afin d'identifier les gènes qui intéragissent avec ZYG-11 pour assumer ses deux fonctions séparables. Par conséquent, nos trouvailles suggèrent un modèle selon lequel ZYG-11 est une sous-unité qui recrute les substrats d'une ligase E3 basée sur CUL-2 qui promeut la progression du cycle cellulaire et empêche l'établissement de la polarité pendant la méiose II, et où le centrosome agit comme la clé qui polarise l'embryon à la fin de la méiose. Summary The mechanisms that couple meiotic cell cycle progression to subsequent developmental events, including specification of embryonic axes, are poorly understood. In the one cell stage embryos of Caenorhabditis elegans, the first signs of Antero-Posterior (A-P) polarity appear after meiosis completion. A centrosome ¬derived component breaks symmetry of the embryo, but the molecular nature of this polarity signal is not known. We established that zyg-11 and cul-2 promote the metaphase to anaphase transition and M phase exit at meiosis II. Our results indicate that ZYG-11 acts as a substrate recruitment subunit of a CUL-2-based E3 ligase. Moreover, we find that the delayed meiosis II exit of embryos lacking zyg-11 is caused by accumulation of the B-type cyclin, CYB-3. We demonstrate that inverted A-P polarity is established during the meiosis II delay in zyg-11(RNAi) and cul¬2(RNAi) embryos. Importantly, we demonstrate that the polarity defects following zyg-11 or cul-2 inactivation can be uncoupled from the cell cycle defects. Furthermore, we found that microtubules appear dispensable for inverted polarity during the meiosis II delay in zyg-11(RNAi) embryos, as well as for A-P polarity during the first mitotic cell cycle in wild-type embryos. We also show the initial results from a comprehensive yeast two hybrid, as well as an RNAi-based functional genomic enhancer and suppressor screen, that may lead to identification of proteins that interact with zyg-11 to ensure the two functions. Our findings suggest a model in which ZYG-11 is a substrate recruitment subunit of an CUL-2-based E3 ligase that promotes cell cycle progression and prevents polarity establishment during meiosis II, and in which the centrosome acts as a cue to polarize the embryo along the AP axis after exit from the meiotic cell cycle.

Sox genes in C. elegans: sox-2 role in postembryonic development

Report for the scientific sojourn carried out at the Columbia University, United States, from 2010 to 2012. Expression of SoxB genes correlates with the commitment of cells to a neural fate; however, the relevance of SoxB proteins in early vertebrate neurogenesis has been difficult to prove genetically due to embryonic lethality and presumed redundant functions. The nematode C. Elegants has only 5 sox genes: sox-2 and sox-3 form the SoxB group while sem-2, sox-4 and egl-13 belong to other Sox groups. Our results show that sox-2 and sem-2 are the sox genes expressed earliest and in a broader manner during embryogenesis, being expressed in several neuronal progenitors. sox-3, sox-4 and egl-13 are expressed in few cells during late embryogenesis, when most neurons are already born. Both sox-2 and sem-2 null mutants are early larval lethal but do not show neuronal specification defects during embryonic development as indicated by quantification of a panneuronal reporter. Potential redundancy or compensatory mechanisms between different sox genes have been ruled out, strongly suggesting that sox genes are not required for specification of embryonically-derived neurons. However, at the first larval stage there are still several blast cells that will give rise to different postembryonic lineages, which generate several neurons amongst other cell types. nterestingly, sox-2 is expressed in many of these progenitor cells. Using mosaic analysis we have so far identified neurons derived from two different postembryonic lineages which fail to be generated in C. elegans sox-2 mutants. These results support the idea that postembryonic progenitor competence is compromised in the absence of sox-2.

Desenvolvimento pré-imaginal de Eueides isabella dianasa (Hübner) (Lepidoptera, Nymphalidae, Heliconiinae) em folhas de Passiflora edulis L. (Passifloraceae)

Lagartas de Eueides isabella dianasa (Hübner, 1806) estão entre os insetos fitófagos que ocorrem em P. edulis. Para esse estudo, foram coletados ovos logo após a oviposição, sendo feitas observações diárias para o registro de eclosão, ecdises (confirmadas pelas cápsulas cefálicas), pupação e emergência de adultos. A viabilidade de ovos foi de 94,9%. O período embrionário observado (em dias ± erro padrão da média) foi de 4,25 ± 0,18. O período larval de 16,9 ± 0.34 dias apresentou cinco ínstares: 2,75 ± 0,2; 2,8 ± 0,2; 3,0 ± 0,1; 3,6 ± 0,2 e 4,6 ± 0,15. O período larval ativo foi de 15,83 ± 0,34 dias, seguido por um período prepupal e pupal de, respectivamente, de 7,42 ± 0,15 (período inativo dentro do desenvolvimento pós-embrionário de 8,4 ± 0,15). O desenvolvimento pós-embrionário foi de 24,25 ± 0,37. A duração do desenvolvimento pré-imaginal de 28,5 ± 0,4 pode ser considerada curta entre os lepidópteros. A duração relativa foi de 14,9% para o período embrionário e de 85,9% para o pós-embironário. O uso do tempo durante o período pós-embrionário foi de 11,3%; 11,7%; 12,4%; 14,8%; 18,9% e 30,6% respectivamente como L1, L2, L3, L4, L5 e pupa. O período larval representou 65,3% enquanto o período inativo (prepupa + pupa) 34,7%. A sobrevivência no final do desenvolvimento, considerando o período de ovo, foi de 27,5% e, considerando o período pós-embrionário, 30,0%, tendo sido mais evidente no quinto ínstar e no período pupal com 63,0% e 64,7%, respectivamente. Este é o primeiro registro de Eueides isabella dianasa no Estado de Alagoas, Brasil.

Visitantes florais do maracujá-amarelo (Passiflora edulis f. flavicarpa Deg. Passifloraceae) em áreas de cultivo com diferentes proximidades a fragmentos florestais na região Norte Fluminense, RJ

Este trabalho teve como objetivos estudar a biologia floral e identificar os principais polinizadores do maracujá-amarelo em áreas de cultivo com diferentes proximidades a fragmentos florestais no norte do estado do Rio de Janeiro. A floração do maracujá-amarelo teve duração de nove meses, no período de setembro a maio. As flores iniciavam a antese às 12:00 h e abriam-se ao longo do dia até às 16:30 h. O processo de curvatura dos estiletes ocorreu ao longo da antese e 72,4% das flores curvaram seus estiletes. A produção de néctar deu-se continuamente, atingindo 18μl/flor/hora e a concentração de solutos totais variou entre 38 e 42%. Xylocopa frontalis e Xylocopa ordinaria foram os principais polinizadores do maracujá-amarelo pela freqüência de visitas, comportamento intrafloral e porte corporal. Estas espécies de abelhas, além de Apis mellifera, estiveram presentes em todas as áreas de cultivo. A maior riqueza de visitantes polinizadores do maracujá-amarelo foi observada em áreas de cultivo próximas a fragmentos florestais, fato relacionado à presença de certos grupos de abelhas nativas, como Centridini e Euglossina (Apidae) que dependem de áreas florestais para nidificação e alimentação. A proximidade a fragmentos florestais também é importante para o fornecimento de recursos alimentares e de nidificação ao longo do ano para a manutenção de populações de Xylocopa.

Distribución y densidad de Percebes Pollicipes elegans (Crustacea: Cirripedia) en el mediolitoral peruano (Yasila, Paita; Chilca, Lima)

Se efectuarón colecciones de "percebes" en diferentes localidades del área comprendida entre Yasila, Paita (05°08'S) y Chilca, Lima (12°29.5'S) identificándose los ejemplares como Pollicipes elegans, determinándose su distribución geográfica, densidad, talla, peso, la relación longitud-peso y algunos bioecológicos. También se recopilo información acerca de su extracción comercial.

Población y biomasa del percebes Pollicipes elegans Lesson, 1830 en las islas Lobos de Tierra y Lobos de Afuera, Perú. 1995-1997

Presenta información sobre el estudio (cálculo y análisis) de la población y biomasa del percebes, realizado desde diciembre 1995 a diciembre 1997 en las áreas mencionadas. Brinda resultados que permiten medir las posibilidades de su explotación racional, considerando las variaciones estacionales y algunas condiciones biológicas.

Evaluación de la población del percebes Pollicipes elegans en las islas Lobos de Afuera (18 - 20 julio 2001)

Presenta los resultados de la evaluación poblacional de percebes Pollicipes elegans en las islas Lobos de Afuera con el fin de estimar la magnitud del stock y determinar las características del recurso en relación al ambiente marino, como base para la toma de decisiones en su ordenamiento pesquero.

Islas Lobos de Afuera: Evaluación de pulpo Octopus mimus Gould, 1852 y percebes Pollicipes elegans (Lesson, 1831), Lambayeque, 2010

La distribución de tallas de pulpo fluctuó entre 40 y 200 mm de longitud del manto. El peso promedio fue 588,20 g, el 85% de los ejemplares no superó el peso mínimo de extracción (1 kg). En hembras predominaron estadios en desarrollo (63%), madurez total (16%); en machos predominaron estadios en desove (64%) y maduros (28%). La mayor concentración del recurso se registró al norte de la isla El Ladrón y frente a isla Quita Calcal. Las tallas de percebes fluctuaron entre 1 y 40 mm de longitud carina-rostral (Lcr), 55% de ejemplares estuvieron maduros. El percebes estuvo distribuido en el intermareal rocoso entre 6°56’9’’S y 6°57’30,6’’S en un área de 887,50 m2. La biomasa se estimó en 3,06 t ±27,18% y la población en 0,42 millones de individuos ±47,30%; el stock de juveniles (<17 mm) fue 0,17 millones de ejemplares; el stock adulto (≥17 mm) estuvo constituido por 0,24 millones de individuos y 2,90 t de la biomasa.

Influência de fungos micorrízicos arbusculares, associada à adição de compostos fenólicos, no crescimento de mudas de maracujazeiro amarelo (Passiflora edulis f. flavicarpus)

Realizou-se um experimento em casa de vegetação, com o objetivo de avaliar a influência da inoculação com fungos micorrízicos arbusculares (FMAs), associada à adição de formononetina (nas concentrações de 0, 5 e 10 µ mol L-1), quercetina e morina (nas concentrações de 0, 5, 10 e 15 µ mol L-1), no crescimento e teor de nutrientes de mudas de maracujazeiro, avaliadas em duas fases: produção das mudas em substrato estéril e após o transplantio para substrato não-estéril. Utilizaram-se as espécies Glomus clarum (Gc) e Glomus fasciculatum (Gf) e uma população nativa de FMAs (IN) isolada de um plantio de maracujá no município de São João da Barra (RJ). Todos os FMAs avaliados (Gc, Gf e IN) proporcionaram aumentos significativos na produção de matéria seca e no teor de nutrientes na fase de produção de mudas e após o transplantio para substrato não-estéril. A aplicação dos compostos fenólicos teve efeito apenas na fase após o transplantio, destacando-se as plantas não inoculadas que mostraram efeito benéfico da aplicação dos flavonóis quercetina e morina e do isoflavonóide formononetina (apenas na concentração 5 µ mol L-1) na colonização radicular pelos FMAs, indicando que tais compostos estimularam a população nativa de FMAs presente no substrato. Nas plantas inoculadas, não se verificou efeito dos compostos na colonização radicular pelo fungo, mas observou-se efeito positivo em algumas das variáveis analisadas.

King Rail Rallus Elegans, 2011

Data sheet produced by the Iowa Department of Natural Resources is about different times of animals, insects, snakes, birds, fish, butterflies, etc. that can be found in Iowa.

Epigenetic regulation of histone H3 serine 10 phosphorylation status by HCF-1 proteins in C. elegans and mammalian cells.

BACKGROUND: The human herpes simplex virus (HSV) host cell factor HCF-1 is a transcriptional coregulator that associates with both histone methyl- and acetyltransferases, and a histone deacetylase and regulates cell proliferation and division. In HSV-infected cells, HCF-1 associates with the viral protein VP16 to promote formation of a multiprotein-DNA transcriptional activator complex. The ability of HCF proteins to stabilize this VP16-induced complex has been conserved in diverse animal species including Drosophila melanogaster and Caenorhabditis elegans suggesting that VP16 targets a conserved cellular function of HCF-1. METHODOLOGY/PRINCIPAL FINDINGS: To investigate the role of HCF proteins in animal development, we have characterized the effects of loss of the HCF-1 homolog in C. elegans, called Ce HCF-1. Two large hcf-1 deletion mutants (pk924 and ok559) are viable but display reduced fertility. Loss of Ce HCF-1 protein at reduced temperatures (e.g., 12 degrees C), however, leads to a high incidence of embryonic lethality and early embryonic mitotic and cytokinetic defects reminiscent of mammalian cell-division defects upon loss of HCF-1 function. Even when viable, however, at normal temperature, mutant embryos display reduced levels of phospho-histone H3 serine 10 (H3S10P), a modification implicated in both transcriptional and mitotic regulation. Mammalian cells with defective HCF-1 also display defects in mitotic H3S10P status. CONCLUSIONS/SIGNIFICANCE: These results suggest that HCF-1 proteins possess conserved roles in the regulation of cell division and mitotic histone phosphorylation.

RAE-1, a novel PHR binding protein, is required for axon termination and synapse formation in Caenorhabditis elegans.

Previous studies in Caenorhabditis elegans showed that RPM-1 (Regulator of Presynaptic Morphology-1) regulates axon termination and synapse formation. To understand the mechanism of how rpm-1 functions, we have used mass spectrometry to identify RPM-1 binding proteins, and have identified RAE-1 (RNA Export protein-1) as an evolutionarily conserved binding partner. We define a RAE-1 binding region in RPM-1, and show that this binding interaction is conserved and also occurs between Rae1 and the human ortholog of RPM-1 called Pam (protein associated with Myc). rae-1 loss of function causes similar axon and synapse defects, and synergizes genetically with two other RPM-1 binding proteins, GLO-4 and FSN-1. Further, we show that RAE-1 colocalizes with RPM-1 in neurons, and that rae-1 functions downstream of rpm-1. These studies establish a novel postmitotic function for rae-1 in neuronal development.