Feedback control of intercellular signalling in development

The intercellular communication that regulates cell fate during animal development must be precisely controlled to avoid dangerous errors. How is this achieved? Recent work has highlighted the importance of positive and negative feedback loops in the dynamic regulation of developmental signalling. These feedback interactions can impart precision, robustness and versatility to intercellular signals. Feedback failure can cause disease.     

Food and metabolic signalling defects in a Caenorhabditis elegans serotonin-synthesis mutant

The functions of serotonin have been assigned through serotonin-receptor-specific drugs and mutants; however, because a constellation of receptors remains when a single receptor subtype is inhibited, the coordinate responses to modulation of serotonin levels may be missed. Here we report the analysis of behavioural and neuroendocrine defects caused by a complete lack of serotonin signalling. Analysis of the C. elegans genome sequence showed that there is a single tryptophan hydroxylase gene (tph-1)-the key enzyme for serotonin biosynthesis. Animals bearing a tph-1 deletion mutation do not synthesize serotonin but are fully viable. The tph-1 mutant shows abnormalities in behaviour and metabolism that are normally coupled with the sensation and ingestion of food: rates of feeding and egg laying are decreased; large amounts of fat are stored; reproductive lifespan is increased; and some animals arrest at the metabolically inactive dauer stage. This metabolic dysregulation is, in part, due to downregulation of transforming growth factor-beta and insulin-like neuroendocrine signals. The action of the C. elegans serotonergic system in metabolic control is similar to mammalian serotonergic input to metabolism and obesity.     

Induction of gut in Caenorhabditis elegans embryos.

Two types of developmental events can cause an embryonic cell to adopt a fate different from that of its neighbours: during a cell division particular contents may be segregated to only one daughter cell and cells may experience different external cues, commonly in the form of inductive cell interactions. Work on development in the nematode Caenorhabditis elegans suggests that most cell fates are specified without a need for cell interactions. In particular, the gut cell lineage of C. elegans has been used as a primary example of specification by differential segregation of determinants. Here I re-examine the role of induction in gut specification by isolating early blastomeres. In C. elegans, the gut derives from all the progeny of a single blastomere (E) of the eight-cell stage. When a gut precursor cell (EMS) is isolated during the first half of the four-cell stage, gut does not differentiate. Gut differentiation is rescued by recombining EMS with its posterior neighbour (P2), but not by recombining EMS with one or both of the other two cells of the four-cell stage. These results demonstrate that P2 induces EMS to form gut in C. elegans.     

A genetic pathway for the development of the Caenorhabditis elegans HSN motor neurons

Thirty-five genes define a pathway for the development of the hermaphrodite-specific neurons (HSNs) in Caenorhabditis elegans. Some of these genes affect only one HSN trait, demonstrating that HSN migration, axonal outgrowth and serotonin expression are mutually independent events in HSN development; others, some of which are regulatory, affect multiple HSN traits. Nearly all are pleiotropic, revealing that the genes specifying HSN development also function in the development of other cell types.     

A blend of small molecules regulates both mating and development in Caenorhabditis elegans

In many organisms, population-density sensing and sexual attraction rely on small-molecule-based signalling systems. In the nematode Caenorhabditis elegans, population density is monitored through specific glycosides of the dideoxysugar ascarylose (the 'ascarosides') that promote entry into an alternative larval stage, the non-feeding and highly persistent dauer stage. In addition, adult C. elegans males are attracted to hermaphrodites by a previously unidentified small-molecule signal. Here we show, by means of combinatorial activity-guided fractionation of the C. elegans metabolome, that the mating signal consists of a synergistic blend of three dauer-inducing ascarosides, which we call ascr#2, ascr#3 and ascr#4. This blend of ascarosides acts as a potent male attractant at very low concentrations, whereas at the higher concentrations required for dauer formation the compounds no longer attract males and instead deter hermaphrodites. The ascarosides ascr#2 and ascr#3 carry different, but overlapping, information, as ascr#3 is more potent as a male attractant than ascr#2, whereas ascr#2 is slightly more potent than ascr#3 in promoting dauer formation. We demonstrate that ascr#2, ascr#3 and ascr#4 are strongly synergistic, and that two types of neuron, the amphid single-ciliated sensory neuron type K (ASK) and the male-specific cephalic companion neuron (CEM), are required for male attraction by ascr#3. On the basis of these results, male attraction and dauer formation in C. elegans appear as alternative behavioural responses to a common set of signalling molecules. The ascaroside signalling system thus connects reproductive and developmental pathways and represents a unique example of structure- and concentration-dependent differential activity of signalling molecules.     

Mobilization of a Drosophila transposon in the Caenorhabditis elegans germ line.

Transposons have been enormously useful for genetic analysis in both Drosophila and bacteria. Mutagenic insertions constitute molecular tags that are used to rapidly clone the mutated gene. Such techniques would be especially advantageous in the nematode Caenorhabditis elegans, as the entire sequence of the genome has been determined. Several different types of endogenous transposons are present in C. elegans, and these can be mobilized in mutator strains (reviewed in ref. 1). Unfortunately, use of these native transposons for regulated transposition in C. elegans is limited. First, all strains contain multiple copies of these transposons and thus new insertions do not provide unique tags. Second, mutator strains tend to activate the transposition of several classes of transposons, so that the type of transposon associated with a particular mutation is not known. Here we demonstrate that the Drosophila mariner element Mos1 can be mobilized in C. elegans. First, efficient mobilization of Mos1 is possible in somatic cells. Second, heritable insertions of the transposon can be generated in the germ line. Third, genes that have been mutated by insertion can be rapidly identified using inverse polymerase chain reaction. Fourth, these insertions can subsequently be remobilized to generate deletion and frameshift mutations by imperfect excision.     

Caenorhabditis elegans has scores of homoeobox-containing genes

Homoeobox-containing genes control cell identities in particular spatial domains, cell lineages, or cell types during the development of Drosophila and Caenorhabditis elegans, and they probably control similar processes in vertebrates. More than 80 genes with homoeoboxes that have sequence similarities ranging from 25 to 100% have been isolated by genetic means or by DNA hybridization to previously isolated genes. We synthesized 500-2,000-fold degenerate oligonucleotides corresponding to a set of well-conserved eight amino acid sequences from the helix-3 region of the homoeodomain. We screened C. elegans genomic libraries with these probes and identified 49 putative homoeobox-containing loci. DNA sequencing confirmed that eight out of ten selected loci had sequences corresponding to the conserved helix-3 region plus additional flanking sequence similarity. One of these genes contained a sequence corresponding to a complete pou-domain and another was closely related to the homoeobox-containing genes caudal/cdx-1. The putative homoeobox loci were mapped to the physical contig map of C. elegans, allowing the identification of potentially corresponding genes from the correlated genetic map. We estimate that the number of homoeobox-containing genes in C. elegans is at least 60, constituting approximately 1% of the estimated total number of genes.     

秀丽隐杆线虫胆固醇7,8-脱氢酶在大肠杆菌中的表达

胆固醇7,8–脱氢酶是参与胆固醇代谢的还原酶,可以将胆固醇转化为7–脱氢胆固醇(合成维生素D3的重要中间体).合成秀丽隐杆线虫的胆固醇7,8-脱氢酶基因daf-36,与具有T7强启动子的载体pET32a(+)连接后,成功构建pET32a-daf-36表达质粒,转入大肠杆菌表达系统E.coli BL21(DE3)和E.coli Rosetta(DE3),经过IPTG诱导培养,蛋白质电泳图谱及质谱结果表明此酶得到了高效表达.     

Chromosomal clustering of muscle-expressed genes in Caenorhabditis elegans

Chromosomes are divided into domains of open chromatin, where genes have the potential to be expressed, and domains of closed chromatin, where genes are not expressed. Classic examples of open chromatin domains include 'puffs' on polytene chromosomes in Drosophila and extended loops from lampbrush chromosomes. If multiple genes were typically expressed together from a single open chromatin domain, the position of co-expressed genes along the chromosomes would appear clustered. To investigate whether co-expressed genes are clustered, we examined the chromosomal positions of the genes expressed in the muscle of Caenorhabditis elegans at the first larval stage. Here we show that co-expressed genes in C. elegans are clustered in groups of 2-5 along the chromosomes, suggesting that expression from a chromatin domain can extend over several genes. These observations reveal a higher-order organization of the structure of the genome, in which the order of the genes along the chromosome id correlated with their expression in specific tissues.     

Control of organ shape by a secreted metalloprotease in the nematode Caenorhabditis elegans.

The molecular controls governing organ shape are poorly understood. In the nematode Caenorhabditis elegans, the gonad acquires a U-shape by the directed migration of a specialized 'leader' cell, which is located at the tip of the growing gonadal 'arm'. The gon-1 gene is essential for gonadal morphogenesis: in gon-1 mutants, no arm elongation occurs and somatic gonadal structures are severely malformed. Here we report that gon-1 encodes a secreted protein with a metalloprotease domain and multiple thrombospondin type-1-like repeats. This motif architecture is typical of a small family of genes that include bovine procollagen I N-protease (P1NP), which cleaves collagen, and murine ADAMTS-1, the expression of which correlates with tumour cell progression. We find that gon-1 is expressed in two sites, leader cells and muscle, and that expression in each site has a unique role in forming the gonad. We speculate that GON-1 controls morphogenesis by remodelling basement membranes and that regulation of its activity is crucial for achieving organ shape.     

Molecular basis of the copulatory plug polymorphism in Caenorhabditis elegans

Heritable variation is the raw material for evolutionary change, and understanding its genetic basis is one of the central problems in modern biology. We investigated the genetic basis of a classic phenotypic dimorphism in the nematode Caenorhabditis elegans. Males from many natural isolates deposit a copulatory plug after mating, whereas males from other natural isolates?including the standard wild-type strain (N2 Bristol) that is used in most research laboratories?do not deposit plugs. The copulatory plug is a gelatinous mass that covers the hermaphrodite vulva, and its deposition decreases the mating success of subsequent males. We show that the plugging polymorphism results from the insertion of a retrotransposon into an exon of a novel mucin-like gene, plg-1, whose product is a major structural component of the copulatory plug. The gene is expressed in a subset of secretory cells of the male somatic gonad, and its loss has no evident effects beyond the loss of male mate-guarding. Although C. elegans descends from an obligate-outcrossing, male?female ancestor, it occurs primarily as self-fertilizing hermaphrodites. The reduced selection on male?male competition associated with the origin of hermaphroditism may have permitted the global spread of a loss-of-function mutation with restricted pleiotropy.     

Transgenerational epigenetic inheritance of longevity in Caenorhabditis elegans

Chromatin modifiers regulate lifespan in several organisms, raising the question of whether changes in chromatin states in the parental generation could be incompletely reprogrammed in the next generation and thereby affect the lifespan of descendants. The histone H3 lysine 4 trimethylation (H3K4me3) complex, composed of ASH-2, WDR-5 and the histone methyltransferase SET-2, regulates Caenorhabditis elegans lifespan. Here we show that deficiencies in the H3K4me3 chromatin modifiers ASH-2, WDR-5 or SET-2 in the parental generation extend the lifespan of descendants up until the third generation. The transgenerational inheritance of lifespan extension by members of the ASH-2 complex is dependent on the H3K4me3 demethylase RBR-2, and requires the presence of a functioning germline in the descendants. Transgenerational inheritance of lifespan is specific for the H3K4me3 methylation complex and is associated with epigenetic changes in gene expression. Thus, manipulation of specific chromatin modifiers only in parents can induce an epigenetic memory of longevity in descendants.     

马拉硫磷对秀丽隐杆线虫的急性毒性研究

研究了马拉硫磷对秀丽隐杆线虫(Caenorhabditis elegans)形态、行为和生理生化等方面的影响,发现马拉硫磷可以导致线虫体型、繁殖能力、衰老、趋化性以及一些代谢酶类变化,但对线虫的寿命、运动能力和靶标酶并未产生显著影响.表明秀丽隐杆线虫对马拉硫磷有较强的耐受性,体内可能存在对马拉硫磷高效的代谢途径.     

Caenorhabditis elegans gene ced-9 protects cells from programmed cell death.

The gene ced-9 of the nematode Caenorhabditis elegans acts to protect cells from programmed cell death. A mutation that abnormally activates ced-9 prevents the cell deaths that occur during normal C. elegans development. Conversely, mutations that inactivate ced-9 cause cells that normally live to undergo programmed cell death; these mutations result in embryonic lethality, indicating that ced-9 function is essential for development. The ced-9 gene functions by negatively regulating the activities of other genes that are required for the process of programmed cell death.     

xnd-1 regulates the global recombination landscape in Caenorhabditis elegans

Meiotic crossover (CO) recombination establishes physical linkages between homologous chromosomes that are required for their proper segregation into developing gametes, and promotes genetic diversity by shuffling genetic material between parental chromosomes. COs require the formation of double strand breaks (DSBs) to create the substrate for strand exchange. DSBs occur in small intervals called hotspots and significant variation in hotspot usage exists between and among individuals. This variation is thought to reflect differences in sequence identity and chromatin structure, DNA topology and/ or chromosome domain organization. Chromosomes show different frequencies of nondisjunction (NDJ), reflecting inherent differences in meiotic crossover control, yet the underlying basis of these differences remains elusive. Here we show that a novel chromatin factor, X non-disjunction factor 1 (xnd-1), is responsible for the global distribution of COs in C. elegans. xnd-1 is also required for formation of double-strand breaks (DSBs) on the X, but surprisingly XND-1 protein is autosomally enriched. We show that xnd-1 functions independently of genes required for X chromosome-specific gene silencing, revealing a novel pathway that distinguishes the X from autosomes in the germ line, and further show that xnd-1 exerts its effects on COs, at least in part, by modulating levels of H2A lysine 5 acetylation.