Assuntos
Eficiência Organizacional , Pesquisa/economia , Pesquisa/organização & administração , Academias e Institutos/organização & administração , Orçamentos , Comércio/métodos , Docentes/organização & administração , Liderança , Apoio à Pesquisa como Assunto , Universidades/organização & administraçãoRESUMO
Proper orientation and positioning of the mitotic spindle is essential for the correct segregation of fate determinants during asymmetric cell division. Although heterotrimeric G proteins and their regulators are essential for spindle positioning in many cell types, their mechanism of action remains unclear. In this study, we show that dyrb-1, which encodes a dynein light chain, provides a functional link between heterotrimeric G protein signaling and dynein activity during spindle positioning in Caenorhabditis elegans. Embryos depleted of dyrb-1 display phenotypes similar to a weak loss of function of dynein activity, indicating that DYRB-1 is a positive regulator of dynein. We find that the depletion of dyrb-1 enhances the spindle positioning defect of weak loss of function alleles of two regulators of G protein signaling, LIN-5 and GPR-1/2, and that DYRB-1 physically associates with these two proteins. These results indicate that dynein activity functions with regulators of G protein signaling to regulate common downstream effectors during spindle positioning in the early C. elegans embryo.
Assuntos
Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/metabolismo , Dineínas/metabolismo , Proteínas Heterotriméricas de Ligação ao GTP/fisiologia , Transdução de Sinais , Fuso Acromático/metabolismo , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/ultraestrutura , Proteínas de Caenorhabditis elegans/antagonistas & inibidores , Proteínas de Caenorhabditis elegans/metabolismo , Dineínas/antagonistas & inibidores , Dineínas/fisiologia , Embrião não Mamífero/metabolismo , Embrião não Mamífero/ultraestrutura , Proteínas de Fluorescência Verde/análise , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Modelos Biológicos , Interferência de RNA , Proteínas Recombinantes de Fusão/análise , Fuso Acromático/ultraestruturaRESUMO
During mitosis different types of cells can have differential requirements for chromosome segregation. We isolated two new alleles of the separation anxiety gene (san). san was previously described in both Drosophila and in humans to be required for centromeric sister chromatid cohesion (Hou et al., 2007; Williams et al., 2003). Our work confirms and expands the observation that san is required in vivo for normal mitosis of different types of somatic cells. In addition, we suggest that san is also important for the correct resolution of chromosomes, implying a more general function of this acetyltransferase. Surprisingly, during oogenesis we cannot detect mitotic defects in germ line cells mutant for san. We hypothesize the female germ line stem cells have differential requirements for mitotic sister chromatid cohesion.
Assuntos
Acetiltransferases/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/enzimologia , Células Germinativas/citologia , Células Germinativas/enzimologia , Mitose , Alelos , Animais , Blastoderma/citologia , Blastoderma/enzimologia , Segregação de Cromossomos , Cromossomos/enzimologia , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Genes de Insetos , Larva/citologia , Larva/enzimologia , Neurônios/citologia , Neurônios/enzimologia , Oogênese , Troca de Cromátide Irmã , Zigoto/citologia , Zigoto/enzimologiaRESUMO
The PAR proteins play an essential role in establishing and maintaining cell polarity. While their function is conserved across species, little is known about their regulators and effectors. Here we report the identification of 13 potential components of the C. elegans PAR polarity pathway, identified in an RNAi-based, systematic screen to find suppressors of par-2(it5ts) lethality. Most of these genes are conserved in other species. Phenotypic analysis of double-mutant animals revealed that some of the suppressors can suppress lethality associated with the strong loss-of-function allele par-2(lw32), indicating that they might impinge on the PAR pathway independently of the PAR-2 protein. One of these is the gene nos-3, which encodes a homolog of Drosophila Nanos. We find that nos-3 suppresses most of the phenotypes associated with loss of par-2 function, including early cell division defects and maternal-effect sterility. Strikingly, while PAR-1 activity was essential in nos-3; par-2 double mutants, its asymmetric localization at the posterior cortex was not restored, suggesting that the function of PAR-1 is independent of its cortical localization. Taken together, our results identify conserved components that regulate PAR protein function and also suggest a role for NOS-3 in PAR protein-dependent cell polarity.
Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Polaridade Celular/genética , Mapeamento Cromossômico/métodos , Genes Supressores , Animais , Animais Geneticamente Modificados , Padronização Corporal/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/fisiologia , Transtornos do Desenvolvimento Sexual , Embrião não Mamífero , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Genes Letais , Masculino , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA/fisiologia , RNA Interferente Pequeno/fisiologia , Proteínas de Ligação a RNA/fisiologia , Supressão Genética , Distribuição TecidualRESUMO
Anillin is a conserved component of the contractile ring that is essential for cytokinesis, and physically interacts with three conserved cleavage furrow proteins, F-actin, myosin II and septins in biochemical assays. We demonstrate that the Drosophila scraps gene, identified as a gene involved in cellularization, encodes Anillin. We characterize defects in cellularization, pole cell formation and cytokinesis in a series of maternal effect and zygotic anillin alleles. Mutations that result in amino acid changes in the C-terminal PH domain of Anillin cause defects in septin recruitment to the furrow canal and contractile ring. These mutations also strongly perturb cellularization, altering the timing and rate of furrow ingression. They cause dramatic vesiculation of new plasma membranes, and destabilize the stalk of cytoplasm that normally connects gastrulating cells to the yolk mass. A mutation closer to the N terminus blocks separation of pole cells with less effect on cellularization, highlighting mechanistic differences between contractile processes. Cumulatively, our data point to an important role for Anillin in scaffolding cleavage furrow components, directly stabilizing intracellular bridges, and indirectly stabilizing newly deposited plasma membrane during cellularization.
Assuntos
Membrana Celular/metabolismo , Proteínas Contráteis/genética , Proteínas Contráteis/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Actinas/metabolismo , Alelos , Sequência de Aminoácidos , Animais , Núcleo Celular/metabolismo , Proteínas Contráteis/química , Citocinese , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , Proteínas de Drosophila/química , Drosophila melanogaster/citologia , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Microscopia Eletrônica de Transmissão , Dados de Sequência Molecular , Mutação/genética , Alinhamento de Sequência , Zigoto/química , Zigoto/metabolismoRESUMO
Caenorhabditis elegans contains a set of six cluster-type homeobox (Hox) genes that are required during larval development. Some of them, but unlike in flies not all of them, are also required during embryogenesis. It has been suggested that the control of the embryonic expression of the worm Hox genes might differ from that of other species by being regulated in a lineal rather than a regional mode. Here, we present a trans-species analysis of the cis-regulatory region of ceh-13, the worm ortholog of the Drosophila labial and the vertebrate Hox1 genes, and find that the molecular mechanisms that regulate its expression may be similar to what has been found in species that follow a regulative, non-cell-autonomous mode of development. We have identified two enhancer fragments that are involved in different aspects of the embryonic ceh-13 expression pattern. We show that important features of comma-stage expression depend on an autoregulatory input that requires ceh-13 and ceh-20 functions. Our data show that the molecular nature of Hox1 class gene autoregulation has been conserved between worms, flies, and vertebrates. The second regulatory sequence is sufficient to drive correct early embryonic expression of ceh-13. Interestingly, this enhancer fragment acts as a response element of the Wnt/WG signaling pathway in Drosophila embryos.