Reverse genetics of Caenorhabditis elegans.

It is somewhat ironic that animals that are the prime choice for detailed genetic analysis, such as the fruit fly and the nematode, have thus far been largely refractory to reverse genetic analysis. Their detailed genetic map, and small genome size have made them subjects of ambitious genome analysis projects, but there is still no strategy to introduce desired changes into their genomes by homologous recombination. Some alternative approaches have recently become available; this review describes possibilities and unsolved problems for reverse genetics in the nematode Caenorhabditis elegans. The transposon Tc1 could prove to be very useful for the isolation of knock out mutants, and possibly also for introduction of more subtle alterations.     

High-throughput reverse genetics: RNAi screens in Caenorhabditis elegans

Two recent chromosome-wide screens for phenotypes caused by RNA-mediated interference (RNAi) in Caenorhabditis elegans have increased our understanding of essential genes in nematodes. These papers represent a major advance in functional genomics.     

The genetics of pathogen avoidance in Caenorhabditis elegans

Much attention is rightly focused on how microbes cause disease, but they can also affect other aspects of host physiology, including behaviour. Indeed, pathogen avoidance behaviours are seen across animal taxa and are probably of major importance in nature. Here, we review what is known about the molecular genetics underlying pathogen avoidance in the nematode Caenorhabditis elegans. In its natural environment, the soil, this animal feeds on microbes and is continuously exposed to a diverse mix of microorganisms. Nematodes that develop efficient behavioural responses that enhance their attraction to sources of food and avoidance of pathogens will have an evolutionary advantage. C. elegans can specifically detect natural products of bacteria, including surfactants (such as serrawettin) and acylated homoserine lactone autoinducers, and it can learn to avoid pathogenic species. To date, several distinct mechanisms have been shown to be involved in pathogen avoidance. They are based on G protein-like, insulin-like and neuronal serotonin signalling. We discuss recent findings on the mechanisms of pathogen recognition in C. elegans, the relationship between alternative behavioural defences and also between these and other life-history traits. We propose that the selective pressure associated with avoidance behaviours influence both pathogen and host evolution.     

Molecular genetics of cell death in the nematode Caenorhabditis elegans

In C. elegans, cell death can be readily studied at the cellular, genetic, and molecular levels. Two types of death have been characterized in this nematode: (1) programmed cell death, which occurs as a normal component in development; and (2) pathological cell death which occurs aberrantly as a consequence of mutation. Analysis of mutations that disrupt programmed cell death in various ways has defined a genetic pathway for programmed cell death which includes genes that perform such functions as the determination of which cells die, the execution of cell death, the engulfment of cell corpses, and the digestion of DNA from dead cells. Molecular analysis is providing insightinto the nature of the molecules that function in these aspects of programmed cell death. Characterization of some genes that mutate to induce abnormal cell death has defined a novel gene family called degenerins that encode putative membrane proteins. Dominant alleles of at least two degenerin genes, mec-4 and deg-1, can cause cellular swelling and late onset neurodegeneration of specific groups of cells. © 1992 John Wiley & Sons, Inc.     

Population genetics of geographically restricted and widespread species of Myrica (Myricaceae)

Allozyme variation of 11 putative loci in five populations of the rare Myrica adenophora Hance, and four populations of its widespread congeneric species, M. rubra (Lour.) Sieb. & Zucc. was studied. Among the 21 alleles studied, no unique allele was detected for M. adenophora, whereas M. rubra had 3 alleles not found in the former species. In terms of genetic diversity, populations of the rare species contained fewer alleles per locus (1.5 versus 1.7), fewer effective number of alleles per locus (1.12 versus 1.20), fewer number of alleles per polymorphic locus (2.14 versus 2.46), lower percentage of polymorphic loci (30.9 versus 40.9), and lower expected heterozygosity (0.106 versus 0.163) than populations of the widespread species. Genetic distances within species average 0.043 for M. adenophora and 0.045 for M. rubra, and between species ranged from 0.052 to 0.177, with a mean of 0.103, which agrees with the very similar gross morphologies of these two species. Intrapopulation differentiation was similar in both species: G(ST) = 0.152 for M. adenophora, and 0.146 for M. rubra, whereas estimated gene flow based on G(ST) values were moderate in these two species (Nm = 1.39 versus 1.46). We inferred that M. rubra and M. adenophora are a progenitor-derivative species pair that emerged before migrating into Taiwan during the last glacial period. We consider the Hengchun population (Chiupeng, Hsuhai, and Chufengpi) and Taitung population (Tienkuan and Lanshan) of M. adenophora which probably arose from two subsets of the genome of M. rubra. Genetic drift was inferred to be one of the forces shaping the observed genetic structure in M. adenophora and M. rubra.     

秀丽隐杆线虫在高校遗传学实验中的应用

秀丽隐杆线虫(Caenorhabditiselegans)是模式生物中的重要成员之一,因其实验成本低,实验周期短,非常适宜用于高校的遗传学实验教学中。线虫在实验教学中的使用,一方面可以有效地丰富高校实验教学的内容,另一方面也可以很好地激发学生的学习兴趣。本文介绍了线虫在遗传学实验教学中的应用实例,如生活周期观察、单因子杂交、单核苷酸多态性研究、RNA干扰(RNAi)实验等;对实验设置、操作要求、实验相关准备工作等进行了较为细致的描述,为线虫在高校遗传学实验教学中的应用提供了详实案例,可为线虫在高校遗传学实验或其他相关实验课程如细胞生物学实验、模式生物与发育生物学实验中的应用提供参考。     

Molecular genetics of cell death in the nematode Caenorhabditis elegans.

In C. elegans, cell death can be readily studied at the cellular, genetic, and molecular levels. Two types of death have been characterized in this nematode: (1) programmed cell death, which occurs as a normal component in development; and (2) pathological cell death, which occurs aberrantly as a consequence of mutation. Analysis of mutations that disrupt programmed cell death in various ways has defined a genetic pathway for programmed cell death which includes genes that perform such functions as the determination of which cells die, the execution of cell death, the engulfment of cell corpses, and the digestion of DNA from dead cells. Molecular analysis is providing insight into the nature of the molecules that function in these aspects of programmed cell death. Characterization of some genes that mutate to induce abnormal cell death has defined a novel gene family called degenerins that encode putative membrane proteins. Dominant alleles of at least two degenerin genes, mec-4 and deg-1, can cause cellular swelling and late onset neurodegeneration of specific groups of cells.     

Recombinational Landscape and Population Genomics of Caenorhabditis elegans

Recombination rate and linkage disequilibrium, the latter a function of population genomic processes, are the critical parameters for mapping by linkage and association, and their patterns in Caenorhabditis elegans are poorly understood. We performed high-density SNP genotyping on a large panel of recombinant inbred advanced intercross lines (RIAILs) of C. elegans to characterize the landscape of recombination and, on a panel of wild strains, to characterize population genomic patterns. We confirmed that C. elegans autosomes exhibit discrete domains of nearly constant recombination rate, and we show, for the first time, that the pattern holds for the X chromosome as well. The terminal domains of each chromosome, spanning about 7% of the genome, exhibit effectively no recombination. The RIAILs exhibit a 5.3-fold expansion of the genetic map. With median marker spacing of 61 kb, they are a powerful resource for mapping quantitative trait loci in C. elegans. Among 125 wild isolates, we identified only 41 distinct haplotypes. The patterns of genotypic similarity suggest that some presumed wild strains are laboratory contaminants. The Hawaiian strain, CB4856, exhibits genetic isolation from the remainder of the global population, whose members exhibit ample evidence of intercrossing and recombining. The population effective recombination rate, estimated from the pattern of linkage disequilibrium, is correlated with the estimated meiotic recombination rate, but its magnitude implies that the effective rate of outcrossing is extremely low, corroborating reports of selection against recombinant genotypes. Despite the low population, effective recombination rate and extensive linkage disequilibrium among chromosomes, which are techniques that account for background levels of genomic similarity, permit association mapping in wild C. elegans strains.     

From cell fates to morphology: developmental genetics of the Caenorhabditis elegans male tail.

The C. elegans male tail is being studied as a model to understand how genes specify the form of multicellular animals. Morphogenesis of the specialized male copulatory organ takes place in the last larval stages during male development. Genetic analysis is facilitated because the structure is not necessary for male viability or for strain propagation. Analysis of developmental mutants, isolated in several functional and morphological screens, has begun to reveal how fates of cells are determined in the cell lineages, and how the specification of cell fates affects the morphology of the structure. Cytological studies in wild type and in mutants have been used to study the mechanism of pattern formation in the tail peripheral nervous system. The ultimate goal is to define the entire pathway leading to the male copulatory organ.     

The genetics of cell migration in Drosophila melanogaster and Caenorhabditis elegans development.

Cell migrations are found throughout the animal kingdom and are among the most dramatic and complex of cellular behaviors. Historically, the mechanics of cell migration have been studied primarily in vitro, where cells can be readily viewed and manipulated. However, genetic approaches in relatively simple model organisms are yielding additional insights into the molecular mechanisms underlying cell movements and their regulation during development. This review will focus on these simple model systems where we understand some of the signaling and receptor molecules that stimulate and guide cell movements. The chemotactic guidance factor encoded by the Caenorhabditis elegans unc-6 locus, whose mammalian homolog is Netrin, is perhaps the best known of the cell migration guidance factors. In addition, receptor tyrosine kinases (RTKs), and FGF receptors in particular, have emerged as key mediators of cell migration in vivo, confirming the importance of molecules that were initially identified and studied in cell culture. Somewhat surprisingly, screens for mutations that affect primordial germ cell migration in Drosophila have revealed that enzymes involved in lipid metabolism play a role in guiding cell migration in vivo, possibly by producing and/or degrading lipid chemoattractants or chemorepellents. Cell adhesion molecules, such as integrins, have been extensively characterized with respect to their contribution to cell migration in vitro and genetic evidence now supports a role for these receptors in certain instances in vivo as well. The role for non-muscle myosin in cell motility was controversial, but has now been demonstrated genetically, at least in some cell types. Currently the best characterized link between membrane receptor signaling and regulation of the actin cytoskeleton is that provided by the Rho family of small GTPases. Members of this family are clearly essential for the migrations of some cells; however, key questions remain concerning how chemoattractant and chemorepellent signals are integrated within the cell and transduced to the cytoskeleton to produce directed cell migration. New types of genetic screens promise to fill in some of these gaps in the near future.     

EMI domains are widespread and reveal the probable orthologs of the Caenorhabditis elegans CED-1 protein

The EMI domain, first named after its presence in proteins of the EMILIN family, was identified here in several metazoan proteins with various domain architectures, among which the mammalian NEU1/NG3 proteins and Caenorhabditis elegans CED-1, identified as a transmembrane receptor that mediates cell corpse engulfment. Functional data available for EMILIN proteins suggest that the EMI domain could be a protein-protein interaction module. Sequence profiles specific of the EMI family of domains led to identify the probable orthologs of the C. elegans CED-1 protein in mammals and insects, which were yet uncovered.     

Population genetics and objectivity in species diagnosis

Species as evolutionary lineages are expected to show greater evolutionary independence from one another than are populations within species. Two measures of evolutionary independence that stem from the study of isolation-with-migration models, one reflecting the amount of gene exchange and one reflecting the time of separation, were drawn from the literature for a large number of pairs of closely related species and pairs of populations within species. Both measures, for gene flow and time, showed broadly overlapping distributions for pairs of species and for pairs of populations within species. Species on average show more time and less gene flow than populations, but the similarity of the distributions argues against there being a qualitative difference associated with species status, as compared to populations. The two measures of evolutionary independence were similarly correlated with F(ST) estimates, which in turn also showed similar distributions for species comparisons relative to population comparisons. The measures of gene flow and separation time were examined for the capacity to discriminate intraspecific differences from interspecific differences. If used together, the two measures could be used to develop an objective (in the sense of being repeatable) measure for species diagnosis.     

Wormnet: a crystal ball for Caenorhabditis elegans

An integrated gene network for Caenorhabditis elegans using data from multiple genome-wide screens encompasses most protein-coding genes and can accurately predict their phenotypes.     

WorfDB: the Caenorhabditis elegans ORFeome Database

WorfDB (Worm ORFeome DataBase; http://worfdb.dfci.harvard.edu) was created to integrate and disseminate the data from the cloning of complete set of approximately 19 000 predicted protein-encoding Open Reading Frames (ORFs) of Caenorhabditis elegans (also referred to as the 'worm ORFeome'). WorfDB serves as a central data repository enabling the scientific community to search for availability and quality of cloned ORFs. So far, ORF sequence tags (OSTs) obtained for all individual clones have allowed exon structure corrections for approximately 3400 ORFs originally predicted by the C. elegans sequencing consortium. In addition, we now have OSTs for approximately 4300 predicted genes for which no ESTs were available. The database contains this OST information along with data pertinent to the cloning process. WorfDB could serve as a model database for other metazoan ORFeome cloning projects.     

Population genetics of the East African White-eye species complex

The Eastern Afromontane biodiversity hotspot consists of isolated mountain massifs embedded within the dry lowland savannas of East Africa and of which the peaks and ridges are covered by cloud forest remnants. These cloud forests are home to the Mountain White-eye (Zosterops poliogaster), while three congeneric species (Abyssinian White-eye, Zosterops abyssinicus; Yellow White-eye, Zosterops senegalensis; Pemba White-eye, Zosterops vaughani) inhabit the adjacent lowland savannas. We sampled individuals of all four species across Kenya to analyse interspecific genetic relationships as well as intraspecific differentiation among mountain populations of Z. poliogaster. While the level of genetic differentiation among the four species was rather low, genetic differentiation within Z. poliogaster was very high, even between geographically neighbouring populations. Overall, levels of genetic variation varied strongly across all four species, with much higher diversity detected within the three lowland ones. The highland species was characterised by numerous private alleles that were geographically restricted at populations from single mountains, some of which showed evidence of recent population bottlenecks. We conclude that Z. poliogaster populations are both of high conservation value and conservation concern, given the high proportion of endemic alleles and the genetic signatures of high genetic drift and low gene flow that are typical for small and isolated populations.