Relative frequencies of different kinds of spontaneous and induced mutants of pneumococci and streptococci capable of growth in the presence of streptomycin

Ravin, Arnold W. (University of Rochester, Rochester, N.Y.), and Ajit K. Mishra. Relative frequencies of different kinds of spontaneous and induced mutants of pneumococci and streptococci capable of growth in the presence of streptomycin. J. Bacteriol. 90:1161-1173. 1965.-Mutations conferring ability to grow in the presence of streptomycin arise spontaneously and can be induced in pneumococci and streptococci. They prove to be of several phenotypic and genetic types, which may be classified as follows: VLR, LR, and HR, which confer resistance, respectively, to less than 50, between 50 and 500, and 500 or more mug/ml of streptomycin. VLR and LR mutations recombine with each other, whereas HR mutations generally replace (do not recombine with) several of the VLR and LR mutations. Spontaneously, the VLR type is several times more frequent than the LR and HR types, which are equally frequent relative to each other. Nitrous acid treatment of deoxyribonucleic acid (DNA) in vitro or ultraviolet irradiation of cells tends to produce the VLR and LR type of mutation. Streptomycin-dependent mutations are rare in the pneumococci and streptococci. One such mutation, requiring 500 to 1,000 mug/ml of streptomycin for optimal growth, arose spontaneously in a group A streptococcal strain, and was transferred via a DNA-mediated transformation to a pneumococcal strain. In the latter, the mutation proved to be very closely linked to the genetic locus at which the streptomycin-resistance mutations arise.     

Interaction of mutations affecting growth rate and resistance to streptomycin in pneumococci and streptococci

Krauss, Marjorie R. (New York University Medical Center, New York, N.Y.), James C. King, and Rody P. Cox. Interaction of mutations affecting growth rate and resistance to streptomycin in pneumococci and streptococci. J. Bacteriol. 92:1337-1344. 1966.-A strain of Streptococcus (Viridans group) was shown by transformation reactions to be the carrier of two interacting mutations. One produced resistance to streptomycin and a slow rate of growth; the only effect of the second was an increase in growth rate when it was added by transformation to streptococcal strains that had already been transformed to bear the first. Similar modifying mutations were observed in strains of streptococci and pneumococci into which the first mutation had been introduced by transformation.     

THE DETECTION OF PLASTICITY GENES IN HETEROGENEOUS ENVIRONMENTS

The molecular genetic mechanisms for phenotypic plasticity across heterogeneous macro- and microenvironments were examined using the Populus genomic map constructed by DNA-based markers. Three hypotheses have been suggested to explain genetic variation in phenotypic response to varying environments (i.e., reaction norm): Lerner's homeostasis, allelic sensitivity, and gene regulation. The homeostasis hypothesis, which predicts that heterozygotes are less sensitive to the environment than homozygotes, was supported for phenotypic plasticity to unpredictable environments (microenvironmental plasticity) at the whole-genome level, but for phenotypic plasticity to predictable environments (macroenvironmental plasticity) the hypothesis was supported only at functioning quantitative trait loci (QTLs). For all growth traits studied, gene regulation was suggested to play a prevailing role in determining the norms of reaction to environments. Indirect evidence for gene regulation is that there tend to be more QTLs with larger effects on the phenotype in optimal growing conditions than suboptimal growing conditions because the expression of these QTLs identified is mediated by regulatory genes. Direct evidence for gene regulation is the identification of some loci that differ from QTLs for trait values within environments and exert an environmentally dependent control over structural gene expression. In this study, fewer environmentally sensitive QTLs were detected that display unparalleled allelic effects across environments. For stem height, there were more regulatory loci and more structural loci (whose expression is determined by gene regulation) affecting phenotypic plasticity than for basal area. It was found that microenvironmental plasticity was likely controlled by different genetic systems than those for macroenvironmental plasticity.     

Nucleolin promotes Ang II‐induced phenotypic transformation of vascular smooth muscle cells by regulating EGF and PDGF‐BB

RNA‐binding properties of nucleolin play a fundamental role in regulating cell growth and proliferation. We have previously shown that nucleolin plays an important regulatory role in the phenotypic transformation of vascular smooth muscle cells (VSMCs) induced by angiotensin II (Ang II). In the present study, we aimed to investigate the molecular mechanism of nucleolin‐mediated phenotypic transformation of VSMCs induced by Ang II. Epidermal growth factor (EGF) and platelet‐derived growth factor (PDGF) inhibitors were used to observe the effect of Ang II on phenotypic transformation of VSMCs. The regulatory role of nucleolin in the phenotypic transformation of VSMCs was identified by nucleolin gene mutation, gene overexpression and RNA interference technology. Moreover, we elucidated the molecular mechanism underlying the regulatory effect of nucleolin on phenotypic transformation of VSMCs. EGF and PDGF‐BB played an important role in the phenotypic transformation of VSMCs induced by Ang II. Nucleolin exerted a positive regulatory effect on the expression and secretion of EGF and PDGF‐BB. In addition, nucleolin could bind to the 5′ untranslated region (UTR) of EGF and PDGF‐BB mRNA, and such binding up‐regulated the stability and expression of EGF and PDGF‐BB mRNA, promoting Ang II‐induced phenotypic transformation of VSMCs.     

THE DIVERGENCE OF NEUTRAL QUANTITATIVE CHARACTERS AMONG PARTIALLY ISOLATED POPULATIONS

The neutral model of phenotypic evolution has yielded several simple predictions about the long-term rates of between-population divergence of polygenic traits and about the equilibrium level of within-population variance when mutation and random genetic drift are the sole evolutionary forces. These conclusions must be modified if populations are only partially isolated. A quantitative model is presented for the development of within- and between-population variance for neutral quantitative characters in pairs of populations with arbitrary effective sizes and migration rates. Both the variance in the base population and subsequent variance generated by mutation are considered, and several dynamical and equilibrium properties are shown to be adequately described by simple approximations. The resultant formulations provide some insight into the sensitivity of measures of morphological distance to gene flow, the necessity of isolation for the accumulation of variation between incipient species, and the consequences of gene flow into captive populations of endangered species.     

IMPLICATION OF INVOLVEMENT OF RAT CHROMOSOME #2 IN SPONTANEOUS TRANSFORMATION OF THE RAT-2 CELL LINE

Using anin vitromodel for cell transformation, the relationship between specific chromosomal aberration and phenotypic changes was studied at different passages of Rat-2 cell line. A marker chromosome resulting from a translocation [t(2;7)] was found to be associated with focus formation in soft agar. Conversely, the loss of this marker chromosome was found to be associated with phenotypic reversion. These results suggest an association of this marker chromosome with phenotypic transformation for the Rat-cell line.     

A mutation in aminopeptidase N (CD13) isolated from a patient suffering from leukemia leads to an arrest in the endoplasmic reticulum

Human aminopeptidase N (APN) is used as a routine marker for myelomonocytic cells in hematopoietic malignant disorders. Its gene and surface expressions are increased in cases of malignant transformation, inflammation, or T cell activation, whereas normal B and resting T cells lack detectable APN protein expression. In this study we elucidated the intracellular distribution, expression pattern, and enzymatic activity of a naturally occurring mutation in the coding region of the APN gene. At physiological temperatures the mutant protein is enzymatically inactive, persists as a mannose-rich polypeptide in the endoplasmic reticulum, and is ultimately degraded by an endoplasmic reticulum-associated degradation pathway. It shows in part the distinct behavior of a temperature-sensitive mutant with a permissive temperature of 32 degrees C, leading to correct sorting of the Golgi compartment accompanied by the acquisition of proper glycosylation but without reaching the cell-surface membrane and without regaining its enzymatic activity. Because the patient bearing this mutation suffered from leukemia, possible links to the pathogenesis of leukemia are discussed.     

QUANTITATIVE VARIATION IN FINITE PARTHENOGENETIC POPULATIONS: WHAT STOPS MULLER'S RATCHET IN THE ABSENCE OF RECOMBINATION?

Finite parthenogenetic populations with high genomic mutation rates accumulate deleterious mutations if back mutations are rare. This mechanism, known as Muller's ratchet, can explain the rarity of parthenogenetic species among so called higher organisms. However, estimates of genomic mutation rates for deleterious alleles and their average effect in the diploid condition in Drosophila suggest that Muller's ratchet should eliminate parthenogenetic insect populations within several hundred generations, provided all mutations are unconditionally deleterious. This fact is inconsistent with the existence of obligatory parthenogenetic insect species. In this paper an analysis of the extent to which compensatory mutations can counter Muller's ratchet is presented. Compensatory mutations are defined as all mutations that compensate for the phenotypic effects of a deleterious mutation. In the case of quantitative traits under stabilizing selection, the rate of compensatory mutations is easily predicted. It is shown that there is a strong analogy between the Muller's ratchet model of Felsenstein (1974) and the quantitative genetic model considered here, except for the frequency of compensatory mutations. If the intensity of stabilizing selection is too small or the mutation rate too high, the optimal genotype becomes extinct and the population mean drifts from the optimum but still reaches a stationary distribution. This distance is essentially the same as predicted for sexually reproducing populations under the same circumstances. Hence, at least in the short run, compensatory mutations for quantitative characters are as effective as recombination in halting the decline of mean fitness otherwise caused by Muller's ratchet. However, it is questionable whether compensatory mutations can prevent Muller's ratchet in the long run because there might be a limit to the capacity of the genome to provide compensatory mutations without eliminating deleterious mutations at least during occasional episodes of sex.     

THE EVOLUTION OF COSTLY MATE PREFERENCES II. THE “HANDICAP” PRINCIPLE

We use a general additive quantitative genetic model to study the evolution of costly female mate choice by the “handicap” principle. Two necessary conditions must be satisfied for costly preference to evolve. The conditions are (i) biased mutation pressure on viability and (ii) a direct relationship between the degree of expression of the male mating character and viability. These two conditions explain the success and failure of previous models of the “handicap” principle. Our model also applies to other sources of fitness variation like migration and host-parasite coevolution, which cause effects equivalent to biased mutation.     

A METHOD FOR THE INVESTIGATION OF ELECTROSTENOLYSIS

A quantitative method for the investigation of electrostenolysis has been developed. Electrostenolysis is redefined in the light of the discovery that organic molecules are subject to it. The experimental requirements for a quantitative study are enumerated, and the apparatus and procedure described. It is found that with ferrous and ferric ions and a cellulose acetate membrane, the potential drop across the membrane must be above about 2150 v./cm. in order to effect any oxidation and reduction. With the present apparatus and conditions the ratio of equivalents oxidized or reduced to faradays passing the membrane is low,— of the order of one to several thousand.     

QUANTITATIVE GENETICS OF GEOMETRIC SHAPE IN THE MOUSE MANDIBLE

Abstract We combine the methods of geometric morphometrics and multivariate quantitative genetics to study the patterns of phenotypic and genetic variation of mandible shape in random‐bred mice. The data are the positions of 11 landmarks on the mandibles of 1241 mice from a parent‐offspring breeding design. We use Procrustes superimposition to extract shape variation and restricted maximum likelihood to estimate the additive genetic and environmental components of variance and covariance. Matrix permutation tests showed that the genetic and phenotypic as well as the genetic and environmental covariance matrices were similar, but not identical. Likewise, principal component analyses revealed correspondence in the patterns of phenotypic and genetic variation. Patterns revealed in these analyses also showed similarities to features previously found in the effects of quantitative trait loci and in the phenotypes generated in gene knockout experiments. We used the multivariate version of the breeder's equation to explore the potential for short‐term response to selection on shape. In general, the correlated response is substantial and regularly exceeds the direct response: Selection applied locally to one landmark usually produces a response in other parts of the mandible as well. Moreover, even selection for shifts of the same landmark in different directions can yield dramatically different responses. These results demonstrate the role of the geometry and anatomical structure of the mandible, which are key determinants of the patterns of the genetic and phenotypic covariance matrices, in molding the potential for adaptive evolution.     

EXPERIMENTAL EVIDENCE FOR THE EVOLUTION OF INDIRECT GENETIC EFFECTS: CHANGES IN THE INTERACTION EFFECT COEFFICIENT,PSI (Ψ), DUE TO SEXUAL SELECTION

Indirect genetics effects (IGEs)—when the genotype of one individual affects the phenotypic expression of a trait in another—may alter evolutionary trajectories beyond that predicted by standard quantitative genetic theory as a consequence of genotypic evolution of the social environment. For IGEs to occur, the trait of interest must respond to one or more indicator traits in interacting conspecifics. In quantitative genetic models of IGEs, these responses (reaction norms) are termed interaction effect coefficients and are represented by the parameter psi (Ψ). The extent to which Ψ exhibits genetic variation within a population, and may therefore itself evolve, is unknown. Using an experimental evolution approach, we provide evidence for a genetic basis to the phenotypic response caused by IGEs on sexual display traits in Drosophila serrata. We show that evolution of the response is affected by sexual but not natural selection when flies adapt to a novel environment. Our results indicate a further mechanism by which IGEs can alter evolutionary trajectories—the evolution of interaction effects themselves.     

Role of the prostanoid FP receptor in action potential generation and phenotypic transformation of NRK fibroblasts

By using an shRNA approach to knockdown the expression of the prostaglandin (PG)-F(2alpha) receptor (FP-R), the role of PGF(2alpha) in the process of phenotypic transformation of normal rat kidney (NRK) fibroblasts has been studied. Our data show that PGF(2alpha) up-regulates Cox-2 expression both at the mRNA and protein level, indicating that activation of FP-R in NRK fibroblasts induces a positive feedback loop in the production PGF(2alpha). Knockdown of FP-R expression fully impaired the ability of PGF(2alpha) to induce a calcium response and subsequent depolarization in NRK cells. However, these cells could still undergo phenotypic transformation when treated with a combination of EGF and retinoic acid, but in contrast to the wild-type cells, this process was not accompanied by a membrane depolarization to -20 mV. Knockdown of FP-R expression also impaired the spontaneous firing of calcium action potentials by density-arrested NRK cells. These data show that a membrane depolarization is not a prerequisite for the acquisition of a transformed phenotype. Furthermore, our data provide the first direct evidence that activity of PGF(2alpha) by putative pacemaker cells underlies the generation of calcium action potentials in NRK monolayers.     

Use of the pneumococcal serovar 3 erythrocyte antibody diagnostic agent for analyzing polysaccharide-containing preparations

Antibody diagnosticum to pneumococci (serovar 3) has been prepared. The analysis of different antigenic preparations of pneumococci (serovars 1,3 and 6B) and the filtrates of culture fluid obtained in the process of the cultivation of bacteria belonging to serovars 3,9N and 23F has been carried out by means of the indirect hemagglutination test. The new diagnosticum has proved to be type-specific. This diagnosticum can be used for the evaluation of the quantitative content of specific polysaccharide in different antigenic preparations of serovar 3 pneumococci, as well as for the determination of the content of specific polysaccharide directly in the culture fluid, which constitutes a step in the determination of the period of the maximum synthesis of polysaccharide, necessary for the optimization of the process of the cultivation of pneumococci.     

CONNECTING QTLS TO THE G-MATRIX OF EVOLUTIONARY QUANTITATIVE GENETICS

Evolutionary quantitative genetics has recently advanced in two distinct streams. Many biologists address evolutionary questions by estimating phenotypic selection and genetic (co)variances ( G matrices). Simultaneously, an increasing number of studies have applied quantitative trait locus (QTL) mapping methods to dissect variation. Both conceptual and practical difficulties have isolated these two foci of quantitative genetics. A conceptual integration follows from the recognition that QTL allele frequencies are the essential variables relating the G -matrix to marker-based mapping experiments. Breeding designs initiated from randomly selected parental genotypes can be used to estimate QTL-specific genetic (co)variances. These statistics appropriately distill allelic variation and provide an explicit population context for QTL mapping estimates. Within this framework, one can parse the G -matrix into a set of mutually exclusive genomic components and ask whether these parts are similar or dissimilar in their respective features, for example the magnitude of phenotypic effects and the extent and nature of pleiotropy. As these features are critical determinants of sustained response to selection, the integration of QTL mapping methods into G -matrix estimation can provide a concrete, genetically based experimental program to investigate the evolutionary potential of natural populations.     

A STUDY OF THE PENETRATION OF MAMMALIAN CELLS BY DEOXYRIBONUCLEIC ACIDS

Tritium-labeled deoxyribonucleic acid (DNA) from pneumococci and from human leukocytes was added to growing cultures of HeLa cells at 37°C. Autoradiography revealed an extensive localization of tritium in the nuclear regions. The label could not be removed by treatment with ribonuclease or dilute perchloric acid, but quantitative removal from the cells could be effected with deoxyribonuclease. Chemical and radioactivity determinations on nucleic acids isolated from the exposed HeLa cells revealed the presence of tritium in all 4 DNA bases. About 12 µg. of tritiated DNA was recovered from 6 x 10⁶ HeLa cells which had been exposed for 24 hours to 240 µg. of the human DNA. From this, it is concluded that the amount of DNA, or its degradation products, taken up by the cells was equivalent to at least 10 per cent of the normal HeLa cell complement.     

ADAPTIVE DIVERGENCE BETWEEN FRESHWATER AND MARINE STICKLEBACKS: INSIGHTS INTO THE ROLE OF PHENOTYPIC PLASTICITY FROM AN INTEGRATED ANALYSIS OF CANDIDATE GENE EXPRESSION

Debate surrounding the integration of phenotypic plasticity within the neo‐Darwinian paradigm has recently intensified, but is largely dominated by conceptual abstractions. Advances in our capacities to identify candidate genes, and quantify their levels of expression, now facilitate the study of natural variation in inherently plastic traits, and may lead to a more concrete understanding of plasticity's role in adaptive evolution. We present data from parapatric threespine stickleback (Gasterosteus aculeatus) demes inhabiting geologically recent, freshwater and saltwater zones of a large estuary. Reaction norms for survival confirm adaptation to local salinity conditions. Analysis of osmoregulatory candidate gene expression within an ecological quantitative genetics framework suggests putative mechanisms underlying adaptive variation, and provides insights into the role of ancestral trait plasticity in this divergence. A sodium–potassium ATPase (ATP1A1) is identified as a candidate gene for freshwater adaptation. In addition to heritable variation for gene expression, we infer significant correlation between measures of expression and individual fitness. Overall results indicate a loss of plasticity in the freshwater deme. We discuss how this is consistent with adaptation facilitated by ancestral plasticity as a heuristic example that may prove useful for future, explicit tests of the genetic assimilation hypothesis.