Identification of bacterial plasmids based on mobility and plasmid population biology

Plasmids contain a backbone of core genes that remains relatively stable for long evolutionary periods, making sense to speak about plasmid species. The identification and characterization of the core genes of a plasmid species has a special relevance in the study of its epidemiology and modes of transmission. Besides, this knowledge will help to unveil the main routes that genes, for example antibiotic resistance (AbR) genes, use to travel from environmental reservoirs to human pathogens. Global dissemination of multiple antibiotic resistances and virulence traits by plasmids is an increasing threat for the treatment of many bacterial infectious diseases. To follow the dissemination of virulence and AbR genes, we need to identify the causative plasmids and follow their path from reservoirs to pathogens. In this review, we discuss how the existing diversity in plasmid genetic structures gives rise to a large diversity in propagation strategies. We would like to propose that, using an identification methodology based on plasmid mobility types, we can follow the propagation routes of most plasmids in Gammaproteobacteria, as well as their cargo genes, in complex ecosystems. Once the dissemination routes are known, designing antidissemination drugs and testing their efficacy will become feasible. We discuss in this review how the existing diversity in plasmid genetic structures gives rise to a large diversity in propagation strategies. We would like to propose that, by using an identification methodology based on plasmid mobility types, we can follow the propagation routes of most plasmids in ?-proteobacteria, as well as their cargo genes, in complex ecosystems.     

The human genome project--some implications of extensive "reverse genetic" medicine

Impressive progress has been made during the past several decades in understanding the pathogenesis of human genetic disease. The tools of molecular biology have allowed the isolation of many disease-related genes by forward and a few by reverse genetics, and the imminent completion of a complete human genetic linkage map will accelerate the genetic characterization of many more genetic diseases. The major impacts of the molecular characterization of human genetic diseases will be 1. To increase markedly the number of human diseases that we recognize to have major genetic components. We already understand that genetic diseases are not rare medical curiosities with negligible societal impact, but rather constitute a wide spectrum of both rare and extremely common diseases responsible for an immense amount of suffering in all human societies. The characterization of the human genome will lead to the identification of genetic factors in many more human diseases, even those that now seem too multifactorial or polygenic for ready understanding. 2. To allow the development of powerful new approaches to diagnosis, detection, screening and even therapy of these disorders aimed directly at the mutant genes rather than at the gene products. This should eventually allow much more accurate and specific management of human genetic disease and the genetic factors in many human maladies. The preparation of a fine-structure physical map of the entire human genome together with an overlapping contiguous set of clones spanning entire chromosomes or large portions of chromosomes is rapidly becoming feasible, and the information that will flow from this effort promises eventually to affect the management of many important genetic diseases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rare but evolutionarily consequential outcrossing in a highly inbred zoonotic parasite

Recurrent self-mating can result in nearly clonal propagation of biological lineages, but even occasional outcrossing can serve to redistribute variation in future generations, providing cohesion among regional populations. The zoonotic parasite Trichinella spiralis has been suspected to undergo frequent inbreeding, resulting in genetically uniform larval cohorts which differ markedly from one another. Here, we explored the extent of inbreeding for this parasite by determining how genetic variation (at variable microsatellite markers) is distributed among 1379 larvae derived from 41 wild boars in Extremadura, Spain. In particular, we sought to determine how much of the genetic variation in this region’s parasites occurs among the larvae of any given wild boar, and whether each derives from one, or more, parental lineages. We found strong evidence for inbreeding, resulting in genetically distinct parasite subpopulations among the parasites derived from many pairs of wild boar. Fully two-thirds of these parasite cohorts appear to derive from inbred parents; in 10% of the wild boars, parasites were so inbred as to become absolutely fixed in all of the assayed genetic loci. In spite of this, more than one pair of parents appear to have given rise to the infections in one-third of the sampled wild boars, resulting in mixed infections. These mixed infections should slow losses of heterozygosity and multi-locus polymorphism in any given parasite lineage. Such outcrossing should limit distinctions that would otherwise accumulate among transmission chains, thereby enforcing cohesion through the region’s population in spite of its marked departure from panmixia. Conditions of transmission may differ in other regions, where such epidemiological features may engender different evolutionary outcomes.     

Understanding the mechanistic regulation of ferroptosis in cancer: the gene matters

Ferroptosis has emerged as a crucial regulated cell death involved in a variety of physiological processes or pathological diseases, such as tumor suppression. Though initially being found from anticancer drug screening and considered not essential as apoptosis for growth and development, numerous studies have demonstrated that ferroptosis is tightly regulated by key genetic pathways and/or genes, including several tumor suppressors and oncogenes. In this review, we introduce the basic concepts of ferroptosis, characterized by the features of non-apoptotic, iron-dependent, and overwhelmed accumulation of lipid peroxides, and the underlying regulated circuits are considered to be pro-ferroptotic pathways. Then, we discuss several established lipid peroxidation defending systems within cells, including SLC7A11/GPX4, FSP1/CoQ, GCH1/BH4, and mitochondria DHODH/CoQ, all of which serve as anti-ferroptotic pathways to prevent ferroptosis. Moreover, we provide a comprehensive summary of the genetic regulation of ferroptosis via targeting the above-mentioned pro-ferroptotic or anti-ferroptotic pathways. The regulation of pro- and anti-ferroptotic pathways gives rise to more specific responses to the tumor cells in a context-dependent manner, highlighting the unceasing study and deeper understanding of mechanistic regulation of ferroptosis for the purpose of applying ferroptosis induction in cancer therapy.     

离子通道与人类遗传病

细胞膜离子通道结构和功能正常是细胞进行生理活动的基础,对离子通道功能具有决定性意义的特定位点的突变导致其开放、关闭或激活、失活功能异常,引起组织机能紊乱,形成各种遗传性疾病。本文从水通道蛋白,钙通道,钠通道,钾通道等多种通道蛋白引起的遗传病的现象以及机理做较深入的阐述。     

人类疾病遗传易感性研究方法进展

遗传易感性是指基于个人遗传背景的多基因遗传病发病风险,即来源于父母一方或双方的特定遗传变异在某些情况下会诱发疾病。在特定疾病的发病机制中某些高外显率的遗传变异发挥重要作用,此类疾病通过患病家系分析即可定位疾病相关遗传变异;但另一些低外显率变异的作用则不明显,需要大规模患病人群分析来解析遗传机制。近年来,随着二代测序和多组学分析技术的发展和基因组数据的大量积累,癌症、代谢性疾病、心脑血管疾病和精神疾病等疾病遗传易感性研究中取得了显著进展,为疾病的早期筛查和诊断治疗提供了参考。     

Biotechnology of the Banana: A Review of Recent Progress

Abstract: A number of biotechnological tools have been developed which could help breeders to evolve new plant types to meet the demand of the food industry in the next century. Available techniques for the transfer of genes could significantly shorten the breeding procedures and overcome some of the agronomic and environmental problems which would otherwise not be possible through conventional methods. In vitro protocols have been standardized to allow commercially viable propagation of desired clones of Musa. An overview of the regeneration of banana by direct and indirect organogenesis, and somatic embryogenesis is presented in this article. In addition, the use of several other biotechnological techniques to enrich the genome of banana, such as selection of somaclonal variants, screening for various useful characteristics, cryopreservation, genetic transformation and molecular genetics are reviewed. In conclusion, the improvement of banana through modern biotechnology should help ensure food security by stabilizing production levels in sustainable cropping systems geared towards meeting domestic and export market demands.     

Sequence alterations can mask each other's presence during screening with SSCP or heteroduplex analysis: BRCA genes as examples

For mutation detection, various screening techniques are widely used because DNA sequencing, the gold-standard method, is still considered to be expensive and laborious for high-throughput screening. Single-strand conformation polymorphism (SSCP) analysis, heteroduplex analysis (HA) and their variant techniques are popular and frequently used for this purpose. It is widely accepted that when searching for unknown sequence variations, any revealed distinct pattern should always be sequenced. We give examples here of the BRCA1 and BRCA2 genes where the SSCP/HA techniques can produce ambiguous predictions if used to detect known genetic variants compared to positive controls. Using direct DNA sequencing, we provide evidence that in such cases, mutations or polymorphisms can mask each other's presence. This phenomenon can often influence the results of any DNA testing because genetic variations such as single-nucleotide polymorphisms occur frequently in the human genome. We suggest that even in the case of known electrophoretic patterns of well-characterized genetic alterations, every sequence alteration should be confirmed by direct DNA sequencing, especially if genetic testing is carried out for diagnostic purposes.     

Using <Emphasis Type="Italic">C. elegans</Emphasis> to screen for targets of ethanol and behavior-altering drugs

Caenorhabditis elegans is an attractive model system for determining the targets of neuroactive compounds. Genetic screens in C. elegans provide a relatively unbiased approach to the identification of genes that are essential for behavioral effects of drugs and neuroactive compounds such as alcohol. Much work in vertebrate systems has identified multiple potential targets of ethanol but which, if any, of those candidates are responsible for the behavioral effects of alcohol is uncertain. Here we provide detailed methodology for a genetic screen for mutants of C. elegans that are resistant to the depressive effects of ethanol on locomotion and for the subsequent behavioral analysis of those mutants. The methods we describe should also be applicable for use in screening for mutants that are resistant or hypersensitive to many neuroactive compounds and for identifying the molecular targets or biochemical pathways mediating drug responses. Published: June 8, 2004.     

Design and implementation of cell-based assays to model human disease.

Cell-based assays, if appropriately designed, can be used to rapidly identify molecular mechanisms of human disease and develop novel therapeutics. In the last 20 years, many genes that cause or contribute to diverse disorders, including cancer and neurodegenerative disease, have been identified. With such genes in hand, scientists have created numerous model systems to dissect the molecular mechanisms of basic cellular and developmental biology. Meanwhile, techniques for high-throughput screening that use large chemical libraries have been developed, as have cDNA and RNA interference libraries that cover the entire human genome. By combining cell-based assays with chemical and genetic screens, we now have vastly improved our ability to dissect molecular mechanisms of disease and to identify therapeutic targets and therapeutic lead compounds. However, cell-based screening systems have yet to yield many fundamental insights into disease pathogenesis, and the development of therapeutic leads is frustratingly slow. This may be due to a failure of such assays to accurately reflect key aspects of pathogenesis. This Review attempts to guide the design of productive cellular models of human disease that may be used in high-throughput chemical and genetic screens. We emphasize two points: (i) model systems should use quantifiable molecular indicators of a pathogenic process, and (ii) small chemical libraries that include molecules with known biological activity and/or acceptable safety profiles are very useful.     

Molecular and chemical genetic approaches to developmental origins of aging and disease in zebrafish

The incidence of diseases increases rapidly with age, accompanied by progressive deteriorations of physiological functions in organisms. Aging-associated diseases are sporadic but mostly inevitable complications arising from senescence. Senescence is often considered the antithesis of early development, but yet there may be factors and mechanisms in common between these two phenomena over the dynamic process of aging. The association between early development and late-onset disease with advancing age is thought to come from a consequence of developmental plasticity, the phenomenon by which one genotype can give rise to a range of physiologically and/or morphologically adaptive states in response to different environmental or genetic perturbations. On the one hand, we hypothesized that the future aging process can be predictive based on adaptivity during the early developmental period. Modulating the thresholds of adaptive plasticity by chemical genetic approaches, we have been investigating whether any relationship exists between the regulatory mechanisms that function in early development and in senescence using the zebrafish (Danio rerio), a small freshwater fish and a useful model animal for genetic studies. We have successfully conducted experiments to isolate zebrafish mutants expressing apparently altered senescence phenotypes during embryogenesis (“embryonic senescence”), subsequently showing shortened lifespan in adulthoods. We anticipate that previously uncharacterized developmental genes may mediate the aging process and play a pivotal role in senescence. On the other hand, unexpected senescence-related genes might also be involved in the early developmental process and regulation. The ease of manipulation using the zebrafish system allows us to conduct an exhaustive exploration of novel genes and small molecular compounds that can be linked to the senescence phenotype, and thereby facilitates searching for the evolutionary and developmental origins of aging in vertebrates. This article is part of a Special Issue entitled: Animal Models of Disease.     

高血压易感基因的分子进化

利用家系连锁分析、候选基因法及全基因组关联研究均未能有效发现普通人群的高血压易感基因或位点。遗传学研究表明, 人类许多疾病易感性的形成与走出非洲时的环境适应性进化密切相关, 这为高血压遗传学研究提供了新思路。文章系统综述了高血压易感基因分子进化研究的理论基础和最新进展, 介绍了本研究小组运用分子进化思路在中国汉族人群高血压遗传学研究中的发现, 对未来的研究方向进行了展望, 以期为高血压和其他疾病的遗传学研究提供参考。     

Contribution of rare and common variants determine complex diseases—Hirschsprung disease as a model

Finding genes for complex diseases has been the goal of many genetic studies. Most of these studies have been successful by searching for genes and mutations in rare familial cases, by screening candidate genes and by performing genome wide association studies. However, only a small fraction of the total genetic risk for these complex genetic diseases can be explained by the identified mutations and associated genetic loci. In this review we focus on Hirschsprung disease (HSCR) as an example of a complex genetic disorder. We describe the genes identified in this congenital malformation and postulate that both common ‘low penetrant’ variants in combination with rare or private ‘high penetrant’ variants determine the risk on HSCR, and likely, on other complex diseases. We also discuss how new technological advances can be used to gain further insights in the genetic background of complex diseases. Finally, we outline a few steps to develop functional assays in order to determine the involvement of these variants in disease development.     

Adaptation, redundancy or resilience

Diversity creates resilience both in ecosystems and living organism. Yet, although genetic diversity protects organisms from many diseases and disorders, it also makes it much harder for geneticist to identify the risk factors that lead to common diseases.The study of the natural environment teaches us that ecological systems rich in biodiversity have greater resilience than less diverse systems, and that resource-poor ecosystems tend to have greater biodiversity to buffer against environmental change. The African savannah, a huge ecosystem, contains an abundance of grasses and other plants, herbivores and their predators. The loss of one species might be compensated for by the presence of others, but if species are relentlessly removed, one after another, the continuing loss will weaken the system until it changes its steady state and eventually collapses.To use another illustrative example of the protection conferred by diversity: modern agriculture uses only six cereal crops as the main basic staples of the human diet. If even one crop were threatened—perhaps by a plant virus or other pathogen—the consequences for humanity would probably be catastrophic. To avoid such a scenario, breeders have created hundreds of cultivars, each with minor phenotypic changes that confer resistance to a biotic or abiotic stressor. Thus, humans too create resilience by increasing biodiversity.In order to improve our understanding of complex diseases, we can extend this notion of diverse ecosystems to organisms. Similarly to the disappearance of one species in an ecosystem with abundant biodiversity, the loss of one gene function might not be immediately apparent, because many such changes can be compensated for, at least partly, by changes in other genes. However, a series of small, cumulative changes in many genes could lead to the breakdown of the phenotype of the organism, rendering it less resilient and more susceptible to disease, especially when it is under environmental or infectious stress. It is like throwing a stone in a pond, which generates small waves; throwing many stones at once causes a more complex disturbance, whereby waves combine to create bigger waves or attenuate each other by interference. Thus, even inherited disorders such as hypertrophic cardiomyopathy show several phenotypes as other genes modify the action of the affected gene.Geneticists have found many genes or whole genomic regions that have multiplied throughout the genome by duplication (Eisenstein, 2010). The repeated sequences might be identical, nearly identical or related, and they can be functional or non-functional, as is the case with pseudogenes. In terms of diversity, repeats have apparently given rise to multigene families, such as the collagens, which encode several structural proteins. Even microorganisms, such as Mycobacterium tuberculosis, have extended gene families or several insertions.It was assumed previously that pseudogenes are unnecessary gene copies and therefore inactivated. Yet, there is increasing evidence that they perform a regulatory role, by influencing the function of the parent gene. The variation in copy number also seems to be as, or even more, important than the number of polymorphisms, particularly in complex diseases or phenotypic traits. One negative example is the gene that codes for glutathione transferase, GSTM1. Roughly half of the population carries a deletion of GSTM1, which reduces their ability to neutralize isothiocyanates. Clearly then, many individuals will have two null alleles and an increased risk of xenobiotic-induced disease. Another fascinating example is that preference for a high-starch diet is associated with multiple copies of the salivary amylase gene, which increases production of this enzyme.Humans show a range of vulnerabilities to complex or infectious diseases, such as pulmonary tuberculosis. Despite an exhaustive search, no obvious, major resistance or susceptibility genes for tuberculosis have been found, although many genes—each with minor effects—have a role in disease susceptibility. Further support for the argument that resilience comes from diversity is found in the confusion around genetic association studies in many complex diseases, in which a given gene might be significantly associated with a condition in one population, but not in others. I suspect that many of these reports can be explained by the fact that susceptibility is caused by cumulative functional changes in many genes along different routes in different groups of humans or animals. In fact, susceptibility to a common disease conferred by a single, major locus would make the organism extremely vulnerable—which is exactly what we see with autosomal-dominant inherited diseases. Thus, it is unlikely that complex diseases are caused by a solitary gene defect, as evolution would select against the high risk of a single dominant effect. Instead, we see a range of conditions and phenotypes, owing to the large number of genes involved.This diversity of genetic factors is a blessing for humanity, as it has equipped us with enormous resilience against many common diseases, from cancer to coronary heart disease, to infectious diseases. But, it is also a bane for the geneticist and the clinical scientists who search for genetic factors that can be used to predict disease susceptibility, or the condition or progress of disease. Complexity and diversity make things far more unpredictable and messy—and therefore more difficult for scientific analysis—but both also ensure our survival against a daily assault of biotic and abiotic stressors.     

Cryptic causation of human disease: reading between the (germ) lines

Most cases of complex human diseases arise sporadically. However, usually there is a significant level of familial aggregation of risk and genetic mapping has identified the responsible gene in a few mendelian cases. Although a disease can be causally genetic, intensified mapping efforts have so far been unable to identify genes that account for more than a small fraction of the familial risk, perhaps because the responsible variation arises by somatic mutation (SM). SM explains the kind of epidemiological pattern seen in cancer, and might have a comparable role in many other diseases. For example, in epilepsy, which has largely defied mapping analysis, the underlying disease pathology, undamped neuronal signaling, is closely connected to gene function. Better technologies to detect and characterize SM are becoming available. However, until it is studied directly, SM will remain a cryptic etiological force, even for diseases that are essentially "genetic".     

Cytokine genes and disease susceptibility

Complex networks of cytokines interact in a dynamic way to homeostatically regulate immune responses and other biological pathways. It is, therefore, not surprising that variation in cytokine level has been correlated with disease susceptibility and process. A fundamental issue is whether such variation is a primary cause for disease or reflects secondary inflammatory change. This can be unravelled by investigating cytokine gene polymorphism to determine whether a genetic basis for cytokine dysregulation is associated with disease. Thousands of disease association studies investigating cytokine gene polymorphisms have been reported although many have not been replicated. This is largely due to lack of statistical power, poor definition of clinical phenotype and lack of matching between cases and controls. An appropriate study design should include: Any genetic analysis of cytokine genes in disease studies should also take into account the fact that cytokines rarely manifest their effects in isolation but rather work in complex regulatory networks. Thus, gene-gene and gene-environment interactions may be at the centre of any disease association. Statistical methods are now being introduced to determine such relationships and this should ultimately allow a more accurate estimate of disease risk for individuals with particular cytokine gene profiles.