Understanding and managing pathogen evolution: a way forward.
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Myotonic dystrophy: will the real gene please step forward!
The mutation underlying myotonic dystrophy (DM) was identified at the end of 1991 amidst great rejoicing from the patients supporting the research and, not least, from those who spent so long searching for it. Subsequently, the molecular genetic phenomena associated with DM have been clearly explained by the transmission behaviour of the expanding repeat, which remains the only mutation that has been described in patients. We understand the molecular basis of anticipation, why the severe congenital form is almost exclusively transmitted by affected mothers and we have widely accepted models of the population genetics of DM. Yet, despite all these clearly explained molecular events, we appear to be hardly any closer to understanding the molecular pathology of DM than when the mutation was first identified. To understand the reason for this, we have to look in detail at the mutation itself, and in particular at the locus and its complex nuances. In doing so, we begin to realise that DM is unique amongst the Mendelianly inherited disorders, in that the mutation, because of its location in a very gene-rich region of the genome, probably simultaneously renders several genes dysfunctional. The somatic heterogeneity of the repeat, coupled with the involvement of several genes, accounts for the pleiotropy observed in the phenotype. Added to this complexity is the uncertainty of the level at which gene dysfunction or gain of function is occurring. It is possibly at the level of DNA/chromatin structure and/or RNA regulation/processing, and all of these pathways may, in different tissues, contribute to the final phenotype.
Animals↗
Locomotor activity responses to ethanol, other alcohols, and GABA-A acting compounds in forward- and reverse-selected FAST and SLOW mouse lines.
Mice selectively bred for high (FAST) or low (SLOW) locomotor stimulant response to ethanol have been found to differ in response to drugs with gamma-aminobutyric acid (GABA)-ergic actions. Reverse selection produced lines that are similar in sensitivity to ethanol stimulation (r-FAST and r-SLOW) and provided a unique model for testing hypotheses about shared genetic influence on sensitivity to ethanol and GABAergic drugs. FAST mice were more stimulated than SLOW mice by all drugs tested: ethanol, methanol, n-propanol, t-butanol, pentobarbital, diazepam, and allopregnanolone. In contrast, r-FAST and r-SLOW mice differed in sensitivity to only a few isolated drug doses. Locomotor responses of each reverse-selected line were significantly different from the responses of their respective forward-selected line for all drugs. Results support an effect of selection for ethanol sensitivity on allosteric modulation of the GABA-A receptor.
Alcohols↗
Effects of heavy metal pollution on genetic variation and cytological disturbances in the Pinus sylvestris L. population.
This isoenzymatic and cytogenetic study has shown significant differences in genetic composition between two groups of Pinus sylvestris trees: tolerant and sensitive to heavy metal pollution. Total and mean numbers of alleles and genotypes per locus were higher in the pollution-sensitive group of trees, but heterozygosity (Ho) was lower in this group. Fixation index (F) indicates that trees tolerant for pollution were in the Hardy-Weinberg equilibrium, while the sensitive group had a significant excess of homozygosity. Cytological analyses demonstrated numerous aberrations of chromosomes in meristematic root tissue of seedlings developed from seeds collected from trees in the polluted area. The aberrations included chromosome bridges and stickiness, laggards, retarded and forward chromosomes, and their fragments. The mitotic index was markedly lower in this group of seedlings, as compared to the control. Both isoenzymatic and cytological analyses showed a significant influence of heavy metal ions on the genetic structure of the Pinus sylvestris population.
Chromosome Aberrations↗
LXIII Cold Spring Harbor Symposium on Quantitative Biology: Mechanisms of Transcription, 3-8 June 1998.
A new perspective is emerging in the transcription field towards understanding gene regulation not only at its most fundamental level but also in the context of chromatin, nuclear compartmentalization, and physiological processes. This direction is being fueled by several key observations. Among them is the discovery of multi-protein complexes whose components reveal a link between gene activity, nuclear structure, and cellular signaling pathways. This information will no doubt be extended by identifying expanded regulatory circuitry using the microchip oligonucleotide array technology. In addition to elucidating the regulatory consequences of these intricate connections, another frontier will be to analyze gene expression within chromosomes. This requires deciphering the mechanism of action of a variety of DNA elements that create a genetic domain such as locus control regions, distal enhancers, insulators, silencers, and matrix attachment regions. Hopefully, with the development of new assays these elements can be as rigorously defined as promoters have been. We can also look forward to capturing critical transcriptional processes by increasingly refined structural analyses. Thus, the scope of problems being addressed in gene regulation has been greatly expanded and the opportunity exists to answer very sophisticated questions in the future.
Archaea↗
Genetic risk for restenosis after coronary balloon angioplasty.
Plain old balloon angioplasty (POBA) is a useful therapeutic strategy especially for angioplasty of small coronary arteries. An association study was performed to identify genes that confer susceptibility to restenosis after POBA. The study population comprised 730 individuals (424 men, 306 women) who underwent successful POBA in at least one major coronary artery and were examined angiographically 6 months after the procedure. A total of 469 subjects (273 men, 196 women) exhibited no restenosis after POBA for any of the coronary lesions, whereas 261 subjects (151 men, 110 women) manifested restenosis for all lesions. The genotypes for 40 polymorphisms of 34 genes were determined with a fluorescence- or colorimetry-based allele-specific DNA primer-probe assay. Multivariate logistic regression analysis with adjustment for age, body mass index, and the prevalence of smoking, hypertension, diabetes mellitus, hypercholesterolemia, and hyperuricemia revealed that two polymorphisms (242C --> T in the NADH/NADPH oxidase p22 phox (p22-PHOX) gene and 2136C --> T in the thrombomodulin (THBD) gene) in men and two polymorphisms (584G --> A in the paraoxonase 1 (PON1) gene and 2445G --> A in the fatty acid-binding protein 2 (FABP2) gene) in women were significantly associated with restenosis after POBA. A stepwise forward selection procedure revealed that the effects of these polymorphisms on restenosis were statistically independent of conventional risk factors for coronary artery disease. Genotyping of these polymorphisms may prove informative for assessment of genetic risk for restenosis after POBA.
Age Distribution↗
Assessment of genetic factors for type 2 diabetes mellitus.
The purpose of the present study was to identify gene polymorphisms for reliable assessment of genetic factors for type 2 diabetes mellitus. The study population comprised 4853 unrelated Japanese individuals (2688 men, 2165 women), including 1489 subjects with type 2 diabetes mellitus (969 men, 520 women) and 3364 controls (1719 men, 1645 women). The genotypes for 148 polymorphisms of 124 candidate genes were determined with a method that combines polymerase chain reaction and sequence-specific oligonucleotide probes with suspension array technology. Sixteen polymorphisms were related (p<0.05) to the prevalence of type 2 diabetes mellitus as determined by the chi-square test. Multivariable logistic regression analysis with adjustment for age, sex, and the prevalence of smoking revealed that, among these polymorphisms, the -603A --> G polymorphism of the gene for coagulation factor III (F3) was significantly (p<0.001) associated with the prevalence of type 2 diabetes mellitus, with the -603G allele representing a risk factor for this condition. A stepwise forward selection procedure demonstrated that F3 genotype (GG versus AA + AG) significantly (p<0.001) and independently affected the prevalence of type 2 diabetes mellitus. Genotype for F3 may prove reliable for assessment of genetic factors for type 2 diabetes mellitus. Determination of the genotype for this gene may contribute to personalized prevention of this condition.
Aged↗
The molecular biology of esophageal adenocarcinoma.
BACKGROUND: Barrett's esophagus is an acquired metaplastic change that occurs in the distal esophagus secondary to chronic gastroesophageal reflux. This premalignant condition forms the most important risk factor for developing esophageal adenocarcinoma, which is an extremely aggressive tumor with a 5-year survival rate of less than 25%. Carcinomas that arise in the setting of Barrett's esophagus are thought to develop as part of the metaplasia-dysplasia-carcinoma sequence. OBJECTIVE: To review the current knowledge on the genomic alterations involved in the development of Barrett's esophagus and its progression to dysplasia and/or cancer. RESULTS: Several changes in gene structure, gene expression, and protein structure are associated with the progression of Barrett's esophagus to adenocarcinoma. Accumulation of these changes seems to be essential, rather than the exact sequence of these changes. Multiple molecular pathways are involved and interact with each other. Alterations in tumor suppressor genes, amongst which p53 and p16, are early events in the metaplasia-dysplasia-adenocarcinoma sequence, followed by loss of cell cycle checkpoints. Ongoing genomic instability leads to cumulative genetic errors and thereby the generation of multiple clones of transformed cells. CONCLUSIONS: Within the multistep process of esophageal adenocarcinogenesis, to date no single molecular marker came forward able to predict who will and who will not develop cancer in the setting of Barrett's esophagus. Instead, panels of markers need to be developed in the future allowing to indicate disease progression. Identification of crucial molecular pathways involved in esophageal adenocarcinogenesis would ultimately improve therapy and facilitate development of new treatment strategies.
Adenocarcinoma↗
Genotraumatic T cells and cutaneous T-cell lymphoma. A causal relationship?
Mycosis fungoides, or cutaneous T-cell lymphoma (CTCL), is a T-cell mediated chronic inflammatory skin disease, which can occasionally progress with a variable time course to a fatal lymphoma or to a leukaemic form called Sézary's syndrome. Extensive research into CTCL has not yet elucidated the primary pathophysiological mechanisms. Immunohistological studies are so far less helpful than expected in establishing early diagnosis and prognosis of the disease. The proposition that an exogenous virus is the cause of CTCL has not been substantiated. Karyotypic analysis of lymphocytes from the skin and blood of patients with CTCL have shown the existence of several genetically aberrant T-cell clones in the same patient. These changes are discussed as potential primary events for the development of CTCL. The hypothesis is put forward that the development of genotraumatic T lymphocytes is involved in the progression of the disease.
Clone Cells↗
A comparison of the CHO/HGPRT+ and the L5178Y/TK+/- mutation assays using suspension treatment and soft agar cloning: results for 10 chemicals.
The mouse lymphoma assay (MLA) and Chinese hamster ovary (CHO) cell assay are sensitive indicators of mutagenicity. The CHO assay has been modified technically to permit treatment in suspension and soft agar cloning comparable to the MLA. This methodology eliminates the risk of metabolic cooperation and the trauma of trypsinization. In addition, a larger population of cells can be treated and cloned for mutant selection. In order to compare the effectiveness of the test systems, 10 chemicals were evaluated for the induction of forward mutations in the CHO and MLA. Several of these chemicals have been reported as clastogenic; therefore, abbreviated colony sizing was performed to gauge the extent of genetic damage to the MLA cells. Both test systems detected benzo[a]pyrene, mitomycin C, acridine orange, and proflavin, and, with the exception of proflavin, more large colonies were present than small colonies. The suspect clastogen, phenytoin, was not mutagenic in the MLA and produced inconclusive results in the CHO. Ethidium bromide, a clastogen and a bacterial mutagen, was not mutagenic in either the MLA or CHO. Four compounds (p-aminophenol, benzoin, methoxychlor, and pyrene) were positive in the MLA, generally inducing a large number of small colonies, while demonstrating no mutagenic activity in the CHO assay. They have also been shown to be generally nongenotoxic in other test systems. Overall, the modified CHO assay did not appear to be better than the MLA for the detection of mutagenic agents. However, the MLA does appear to have lower specificity.
Agar↗
Posttraumatic stress disorder: a sensitization reaction.
This article discusses past research bearing on the question of the etiology of Posttraumatic Stress Disorder (PTSD). It argues that PTSD can be adequately accounted for by a process of emotional sensitization and that this is a more parsimonious explanation than the two-factor learning theory of Mowrer, now postulated by several writers. In brief, the etiology and subsequent development of PTSD is viewed as the result of the sensitization of fear/anxiety which is linked to a variety of to be conditional stimuli by both backward and forward association: these become conditional stimuli (CSi) once paired with the instigating circumstances. It is furthermore assumed that PTSD will not occur in the absence of a genetic susceptibility that may vary from zero to absolute certainty. Thus far, our evidence is limited to a sensitivity to loud sounds, but it is highly probable that touch and other sensory systems are involved (not necessarily in parallel). The fact that abuse often leads to behavioral disorders, including sexually seductive behaviors in children sexually abused, requires a recognition that emotional reactions other than fear may be sensitized. Fear in combination with pleasure or pleasure alone coupled with a loss of self-esteem may explain these acting-out behaviors.
Adult↗
The drugs don't work-or do they? Pharmacological and transgenic studies of the contribution of NMDA and GluR-A-containing AMPA receptors to hippocampal-dependent memory.
OBJECTIVE: The aim of this article is to provide a review of studies using N-methyl-D-aspartate (NMDA) receptor antagonists to assess the hippocampal long-term potentiation (LTP)/learning hypothesis. DISCUSSION: In particular, we will re-examine the validity of both (1) the original hippocampal LTP/spatial learning hypothesis of Morris and (2) the sensorimotor account put forward by Cain, among others, both from the point of view of the pharmacological studies on which they were based and with regard to recent studies with genetically modified mice. More specifically, we will review the pharmacological studies in the light of recent work on the glutamate receptor A (GluR-A or GluR1) L-alpha-amino-3-hydroxy-5-methyl-4-isoxazelopropionate (AMPA) receptor sub-unit knockout mouse. We will argue that neither the original hippocampal LTP/spatial learning hypothesis nor a sensorimotor account can adequately explain all of the available data. We argue instead that hippocampal synaptic plasticity, which requires NMDA receptors for its induction and GluR-A-containing AMPA receptors for its continued expression, contributes to a process whereby appropriate behavioural responses are selected rapidly on the basis of conditional information provided by the context. These contextual cues could include not only the spatial context (i.e. the 'where') and the temporal context (the 'when'), but also other aspects of context, such as internal state cues (hunger and fear state), and can be used to rapidly and flexibly alter valences of specific response options. RECOMMENDATIONS: We also suggest that there is a separate, distinct, NMDA/GluR-A-independent mechanism through which the context can gradually (incrementally or decrementally) alter the valence of a particular response option.
2-Amino-5-phosphonovalerate↗
A review of physiological prestress adaptation: effects of duration and context.
This review focuses on 3 aspects of experimental designs for prestress, physiological adaptation periods: (1) effects of duration on stabilization of various physiological parameters:(2) effect of type of activity and instruction on prestress physiological adaptation: and (3) definitions of physiological baselines. Analyses of the literature revealed that for many physiological parameters it remains unclear how much time is required for stabilization and that for the remaining variables the necessary duration of prestress adaptation varies greatly, with blood pressure potentially being the slowest variable to stabilize. Effects of type of activity are essentially unknown. Instructions which create certainly about experimental procedures in subjects tend to facilitate adaptation to the laboratory environment. Baseline definitions were remarkably variable with no less than 15 different definitions being used across the 50 studies reviewed. Suggestions for future research and recommendations for prestress experimental designs are put forward in the discussion.
Adaptation, Physiological↗
Plasmacytoid dendritic cell-mediated L-glutamate catabolism links gut microbiota to male infertility.
Emerging evidence suggests that gut microbiota composition influences male reproductive health; however, the immunometabolic mechanisms underlying this association remain insufficiently characterized. We investigated whether specific immune cell-mediated metabolic pathways, particularly plasmacytoid dendritic cell (pDC)-driven L-glutamate catabolism via the hydroxyglutarate pathway, contribute to the causal link between gut microbiota and male infertility. We conducted a 2-sample, 2-step Mendelian randomization (MR) analysis using inverse-variance weighting as the primary estimator and Bayesian weighted MR for robustness. Exposure data comprised 412 gut microbial taxa/metabolic pathways and 731 immune cell phenotypes from large European-ancestry genome-wide association studies. Male infertility genome-wide association studies data (1429 cases; 128,710 controls) were obtained from FinnGen R10. Only exposure-mediator-outcome pairs meeting stringent pleiotropy, heterogeneity, and reverse-causality criteria were retained for mediation analysis. Nine microbial taxa/metabolic pathways and 18 immune traits exhibited putative causal associations with male infertility. The L-glutamate degradation V pathway via hydroxyglutarate was linked to reduced infertility risk (inverse-variance weighting odds ratio [OR] = 0.68; 95% confidence interval, 0.52-0.89; P = .005). Two-step MR suggested that forward scatter area on pDCs may mediate this association, although the mediation effect was imprecise (effect = 0.0277; 95% confidence interval, -0.0348 to 0.0903). This study provides suggestive genetic evidence that pDC-mediated glutamate catabolism may connect gut microbial metabolic activity to male infertility. These findings highlight immunometabolic pathways as testable targets for mechanistic validation and microbiota-directed interventions.
Male↗
Role of homocysteine synthetase in an alternate route for methionine biosynthesis in Saccharomyces cerevisiae.
In vivo studies have shown that, in the absence of homoserine-O-transacetylase activity (locus met(2)), the C(4)-carbon moiety of ethionine is utilized (provided the ethionine resistance gene eth-2r is present) by methionine auxotrophs, except for met(8) mutants (homocysteine synthetase-deficient). Concomitant utilization of sulfur and methyl group from methylmercaptan or S-methylcysteine has been demonstrated. In the absence of added methylated intermediates, the methyl group of methionine formed from ethionine is derived from serine. In vitro studies with crude extracts of Saccharomyces cerevisiae have demonstrated that this synthesis of methionine occurs by the following reactions: CH(3)-SH + ethionine right harpoon over left harpoon methionine + C(2)H(5)SH and S-methylcysteine + ethionine right harpoon over left harpoon methionine + S-ethylcysteine. In the forward direction, the second product of the second reaction was shown to be S-ethylcysteine; this reaction has also been found reversible, leading to ethionine formation. Genetic and kinetic data have shown that homocysteine synthetase catalyzes these two reactions, at 0.3% of the rate it catalyzes direct homocysteine synthesis: O-Ac-homoserine + Na(2)S --> homocysteine + acetate. The three reactions are lost together in a met(8) mutant and are recovered to the same extent in spontaneous prototrophic revertants from this strain. Methionine-mediated regulation of enzyme synthesis affects the three activities and is modified to the same extent by the presence of the recessive allele (eth-2r) of the regulatory gene eth-2. Affinities of the enzyme for substrates of both types of reactions are of the same order of magnitude. Moreover, ethionine, the substrate of the second reaction, inhibits the third reaction, whereas O-acetyl-homoserine, the substrate of the third reaction, inhibits the second reaction. An enzymatic cleavage of S-methylcysteine, leading to methylmercaptan production, has been shown to occur in crude yeast extracts. It is concluded that the enzyme homocysteine synthetase participates in the two alternate pathways leading to methionine biosynthesis in S. cerevisiae, one involving O-acetyl-homoserine and H(2)S, the other involving the 4-carbon chain of ethionine and a mercaptyl donor. Participation of the two types of reactions catalyzed by homocysteine synthetase, in in vivo methionine synthesis, has been shown to occur in a met(2) partial revertant.
Amino Acids↗
Accumulation of arginine precursors in Escherichia coli: effects on growth, enzyme repression, and application to the forward selection of arginine auxotrophs.
The accumulation or ornithine, citrulline, and possibly acetylornithine by Escherichia coli K-12 arginineless mutants provided with acetylarginine as source of arginine causes severe growth inhibition. This occurs under conditions where comparable derivatives of E. coli W (Bollon and Vogel, 1973) show little or no growth inhibition. The same conditions, which have been reported to cause noncorrelative synthesis of acetylornithinase and argininosuccinase in E. coli W (Bollon and Vogel, 1973), do not alter the correlative pattern of enzyme synthesis observed in E. coli K-12. Moreover, previously reported effects of ornithine and citrulline on repression of the arginine regulon in E. coli W are not observed in the K-12 strains examined. The bearing of these observations on possible differences between the mechanism of enzyme repression operating in the two types of strains cannot yet be fully evaluated; it is, however, clear that considerable care should be exercised before extrapolating the results obtained with one type of strain to the other one. The particularly strong inhibition of acetylarginine utilization exerted by ornithine in E. coli K-12 allows the forward selection of several classes of arginine auxotrophs from strains deficient in carbamoylphosphate biosynthesis and thus capable of ornithine accumulation. Possible applications of this technique to the genetic analysis of the bipolar argECBH operon are discussed.
Arginine↗
The results of assays in Drosophila as indicators of exposure to carcinogens.
Drosophila has fulfilled a dual function in the field of genetic toxicology: for use in short-term tests for identifying carcinogens and in a model for studies of the mechanisms of mutagenesis by chemicals. Until the mid-1980s, use of Drosophila in short-term tests was restricted to assays for genetic damage in germ cells, mostly in males. The largest database, on 700-750 chemicals, is available for the test for sex-linked recessive lethal (SLRL) forward mutation. The database for assays of the consequences of chromosomal breakage--reciprocal translocations and chromosome loss--is smaller, with about 100 chemicals tested. Comparative studies conducted within the US National Toxicology Program showed that SLRL is a better end-point than reciprocal translocation: of 66 chemicals (68 entries) that induced SLRL, only 28 (41%) induced reciprocal translocation. The major weakness of the SLRL assay is its low sensitivity (0.27-0.79) for mammalian genotoxins. A strength of the SLRL mutation test is its high specificity, which is close to 1. Thus, whereas a negative response in Drosophila provides little evidence for genotoxicity, a positive response (SLRL frequency > or = five times the control level) provides good evidence that a chemical is a trans-species mutagen and probably also carcinogenic to mammals. The poor performance of the SLRL test revealed in several collaborative studies led to the development of assays for recombination in somatic cells of Drosophila. Two of these tests have been evaluated for all known classes of genotoxic chemical: the mwh/flr wing spot test on more than 400 chemicals and the white/white+ eye spot test on about 220 chemicals. Of 24 carcinogens that gave negative or inconclusive test results in the SLRL assay, 22 gave positive results in one or both of the somatic systems. Their better performance in comparison with the germ-line assays is primarily the result of their low cost (5-10% of that needed for an SLRL assay), allowing use of multiple doses and protocols and the use of distinct tester strains with heterogeneity for activation of procarcinogens. For qualitative and quantitative studies on structure-activity and activity-activity relationship, only germ-line system have been used. In general, clear relationships between physico-chemical parameters (s values, O6/N7-alkylguanine ratios), carcinogenic potency in rodents and several descriptors of genotoxic activity in germ cells (from mice and Drosophila) became apparent when the following descriptors were used: (1) estimates of TD50 (lifetime doses expressed in milligrams per kilogram body weight or millimoles per kilogram body weight) from bioassays for cancer in rodents; (2) the degree of germ-cell specificity, i.e. the ability of a genotoxic agent to induce mutations at practically any stage of development of Drosophila and mouse spermatogenesis, as opposed to a more specific response in postmeiotic stages of both species; (3) the M(NER-)/M(NER+) hypermutability ratio, determined in a repair assay in Drosophila germ cells; (4) the ratio of chromosomal aberrations to SLRL in postmeiotic germ cells of Drosophila, i.e. the comparative efficiency of a carcinogen to induce these two end-points; (5) mutational spectra induced at single loci, i.e. the seven loci used in the specific-locus test in mice and the vermilion, white and rosy genes of Drosophila; and (6) the doubling doses in milligrams or millimoles per kilogram for specific locus induction in mice. On the basis of these parameters, alkylating agents were classified into three categories in terms of germ-cell specificity, which is primarily due to stage-related differences in DNA repair, clastogenic efficiency, type of mutation spectra and carcinogenic potency in rodents. The three categories allow predictions of the genotoxicity of alkylating agents but not yet for other categories of genotoxic carcinogens.
Animals↗
Heterozygous alpha 1-antitrypsin deficiency: a longitudinal lung function study.
alpha 1-antitrypsin phenotypes were determined in 906 adults from an entire community; 4.9 percent were heterozygotes (Pi MZ) for severe deficiency and 8 percent were heterozygotes (Pi MS) for mild deficiency. Lung function both in cross-section and over a period of three years was examined in those subjects with the two heterozygote deficiency phenotypes and compared with the lung function of those with the normal Pi M phenotype. FVC and FEV1 were standardised for age, sex, height, race, smoking habit and quantum, and respiratory symptoms. Using techniques of multivariate analysis and rigorous matched pairs there was no evidence of a deterioration in lung function in Pi MS or Pi MZ subjects. The tests used did not exclude the possibility of minor alveolar abnormalities but the results do allow the confident counselling of heterozygotes that they can look forward to full respiratory health.
Adult↗