A “chromosomic” recombination theory for multiplicity reactivation in phages

Summary With the assumption of inactivation of small traits in bacteriophages chromosomes by ultraviolet irradiation the probability of multiplicity reactivation of irradiated phages is calculated. The result appears to be in agreement with the experimental results ofDulbecco.In the mathematical treatment of the problem a distinction is made between ordinary genes, with probability of inactivation negligible relative to the probability of inactivation of the whole phage, and a few vulnerable centers or genes whose probability of inactivation is not negligible. The hypothesis of the existence of vulnerable centers appears necessary in order to explainDulbecco's experimental results.

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Zusammenfassung Die Wahrscheinlichkeit der Reaktivation durch multiplen Infektionen (multiplicity reactivation) ist unter Voraussetzung, dass die Ultraviolett-Bestrahlung kleine Teile der Bakteriophagenchromosomen inaktiviert, berechnet. Das Ergebnis entspricht den experimentellen ResultatenDulbecco's.In der mathematischen Behandlung des Problems ist ein Unterschied gemacht worden zwischen gewöhnlichen Genen — wovon jeder eine geringe Wahrscheinlichkeit der Inaktivierung hat, in Bezug auf die Wahrscheinlichkeit der Inaktivierung der ganzen Bakteriophage — und einige verletzbare Stellen oder Genen, wovon die Wahrscheinlichkeit der Inaktivierung grössenordnungsmÄssig höher liegt. Die Hypothese über die verletzbaren Stellen war notwendig um die experimentellen ResultateDulbecco's zu erklÄren.

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Résumé Postulant que de petites sections des chromosomes du bactériophage sont inactivés par l'irradiation ultraviolette, on a calculé la probabilité de réactivation par infection multiple (multiplicity reactivation) des bactériophages traités par les rayons ultraviolets. Les résultats obtenus s'accordent avec les résultats expérimentaux de M.Dulbecco.Dans la tractation mathématique des problèmes une distinction a été faite entre les gènes ordinaires — dont la probabilité d'inactivation prise individuellement peut Être négligée relativement à la probabilité d'inactivation du phage entier — et quelques centres vulnérables ou gènes dont la probabilité d'inactivation ne pourrait Être négligée.L'hypothèse de l'existance des centres vulnérables s'est démontrée nécessaire à l'explication des résultats expérimentaux de M.Dulbecco.

     

Origin of genetic variation: regulation of genetic recombination in the higher organisms — a theory

Summary Recent studies in the fungi, particularly Neurospora and Schizophyllum, have revealed a number of genetic features which, viewed in conjunction with earlier observations on other organisms, form a pattern, or model, which appears to be basic to the control of recombination in all eukaryotes, including higher organisms. It is assumed that the control is exercised on mechanisms that produce new alleles through recombination, as understood in broad terms and including such a likely phenomenon as gene conversion, which may or may not involve crossing-over, as well as equal and unequal crossing-over. The recombination may thus occur between alleles in either the homozygous or heterozygous condition. In the model, regulatory genes and breeding behaviour are integrated into one self-regulatory system controlling the production of new genetic variation.The model is based on the following five general features, largely substantiated by the results in Neurospora and Schizophyllum: 1) The frequency of recombination in a particular chromosomal region is controlled by specific regulatory genes (rec). 2) There may be a number of such specific, regulatory genes responsible for recombination in a given region. 3) A rec. locus may influence recombination in more than one region. 4) The regulatory genes have no specific physical relationship with the region(s) they control, and are usually located at random in the genome. 5) Of the allelic forms of the regulatory genes it is always the dominant gene which suppresses recombination and the recessive gene which increases recombination. The rec system is epistatic to other genetic elements jointly involved in the overall control of recombination in a specific region. It is suggested that usually the control of recombination in a given region is exercised, cumulatively, by the balance of the dominant and recessive genes of the specific rec loci in the organism. Outbreeding, with the associated high heterozygosity of the regulatory rec loci, virtually switches off recombination, producing few new variations. Inbreeding produces homozygosity of these loci, resulting in certain individuals which will have a considerable number of their regulatory loci in the homozygous recessive condition and in which recombination will be switched on, producing new variation at a high frequency. Inbreeding is thus an integrated, evolutionary system of considerable importance, and is not a degenerate dead end, as many investigators have previously thought.The model has another compensatory function in evolution. In major loci, or in an operon, where there are structural genes and closely linked operator genes, as exemplified by the S locus, there are indications that the present model is concerned with the regulation of both structural and operator genes. The consequences of the model in the two classes of genes, however, are in direct contrast to each other: High heterozygosity which is instrumental in switching off recombination, and which is therefore helpful in maintaining stability in the structural gene, is conducive to functional variation of the operator gene; and high homozygosity, which is instrumental in switching on recombination, and which is therefore helpful in producing variation in the structural gene, is conducive to the stability of the operator gene.This model of the control of genetic variation in a specific chromosomal region is significant in development as well as in evolution, and throws light on a number of hitherto intractable problems peculiar to the higher organisms. For example, the model is helpful in explaining: 1) the origin of new self-incompatibility alleles in the flowering plants; 2) the impressive speciation in the waif flora (and fauna) of the oceanic islands; 3) the presence of high genetic variability in inbreeding species of plants; 4) environmentally-induced heritable variation in certain plants; and 5) the genetic mechanism of antibody diversity in animals.     

Integration of function in the nervous system — A new theory

A new theory of synaptic function in the nervous system (Dempsher, 1978) is applied to the simplest system for integration of function in the nervous system. This system includes a sensory and motor neuron and three ‘synaptic’ regions associated with those two neurons; a receptor region, an interneuronal spinal synaptic region linking the two neurons, and an effector region. Information is first received and processed at the receptor region. The processing consists of five components:

1. A highly selective mechanism which allows only that information to enter the receptor system which is appropriate.
2. The ‘appropriateness’ of the information is determined by the alphabet (miniature potentials) already in that area.
3. The information entering the system is assembled in a pattern meaningful for the next processing operation.
4. The assembled information is then ‘disassembled’ into its subunits and mapped into the alphabet (miniature potentials).
5. These miniature potentials are assembled into another pattern meaningful to fit the role of the receptor region.
6. This new pattern is repacked for transit to the central synaptic region.

At the central synaptic region, essentially the same process takes place except here an additional operation takes place which determines its role in the processing system. The incoming information is disassembled into its subunits, mapped into the miniature potentials already there; these are collected together in a meaningful pattern, ‘operated’ on, then repacked for transit to the effector site, where again the same kind of processing sequence takes place. In all three regions, despite the difference in their roles, there are similar processing features:

1. In each region, three forms of the nerve impulse are involved: miniature graded potentials, graded potentials, action potentials.
2. In each region, each component of the process is carried out by a precise mathematical operation: four each in the receptor and effector regions; five in the central synaptic region.

It is suggested that integration of function in the nervous system consists of converting information into energy which is in turn converted into a number. Processing of information at each region then involves mathematical operations applied to these numbers. Function appears to be stereotyped in all three regions. The receptor region receives highly selective and restrictive information so that the universe we ‘perceive’ would appear to be a subset of a much larger universe.     

High efficiency versus maximal performance — The cause of oxidative stress in eukaryotes: A hypothesis

Degenerative diseases are in part based on elevated production of ROS (reactive oxygen species) in mitochondria, mainly during stress and excessive work under stress (strenuous exercise). The production of ROS increases with increasing mitochondrial membrane potential (ΔΨ_m). A mechanism is described which is suggested to keep ΔΨ_m at low values under normal conditions thus preventing ROS formation, but is switched off under stress and excessive work to maximize the rate of ATP synthesis, accompanied by decreased efficiency. Low ΔΨ_m and low ROS production are suggested to occur by inhibition of respiration at high [ATP]/[ADP] ratios. The nucleotides interact with phosphorylated cytochrome c oxidase (COX), representing the step with the highest flux-control coefficient of mitochondrial respiration. At stress and excessive work neural signals are suggested to dephosphorylate the enzyme and abolish the control of COX activity (respiration) by the [ATP]/[ADP] ratio with consequent increase of ΔΨ_m and ROS production. The control of COX by the [ATP]/[ADP] ratio, in addition, is proposed to increase the efficiency of ATP production via a third proton pumping pathway, identified in eukaryotic but not in prokaryotic COX. We conclude that ‘oxidative stress’ occurs when the control of COX activity by the [ATP]/[ADP] ratio is switched off via neural signals.     

Mus81 cleavage of Holliday junctions: a failsafe for processing meiotic recombination intermediates?

The Holliday junction (HJ) is a central intermediate of homologous recombination. Its cleavage is critical for the formation of crossover recombinants during meiosis, which in turn helps to establish chiasmata and promote genetic diversity. Enzymes that cleave HJs, called HJ resolvases, have been identified in all domains of life except eukaryotic nuclei. Controversially, the Mus81-Eme1 endonuclease has been proposed to be an example of a eukaryotic nuclear resolvase. However, hitherto little or no HJ cleavage has been detected in recombinant preparations of Mus81-Eme1. Here, we report the purification of active forms of recombinant Schizosaccharomyces pombe Mus81-Eme1 and Saccharomyces cerevisiae Mus81-Mms4, which display robust HJ cleavage in vitro, which, in the case of Mus81-Eme1, is as good as the archetypal HJ resolvase RuvC in single turnover kinetic analysis. We also present genetic evidence that suggests that this activity might be utilised as a back-up to Mus81-Eme1's main activity of cleaving nicked HJs during meiosis in S. pombe.     

Are there morphologically abnormal early recombination nodules in the Drosophila melanogaster meiotic mutant mei-218?

Early recombination nodules have been suggested to perform a role in meiotic gene conversion recombination events. The meiotic recombination-defective mutant mei-218 greatly reduces the frequency of meiotic crossover (reciprocal) recombination events and reduces the number of late recombination nodules to the same extent. However, it does not reduce the frequency of simple gene conversion events, although they are abnormal in having shorter coconversion tracts than controls. The original cytological study yielded somewhat fewer early nodules in mei-218 than in controls, although very abnormal ones might have been missed. The present study failed to identify a mei-218 specific abnormal category. However, because recombination nodules are at present recognizable only by their morphology, a definitive answer to this question must await a specific probe for recombination nodules. Moreover, the possibility remains that early nodules in mei-218 are more ephemeral than are early nodules in wild type.     

The spatial regulation of meiotic recombination hotspots: Are all DSB hotspots crossover hotspots?

A key step for the success of meiosis is programmed homologous recombination, during which crossovers, or exchange of chromosome arms, take place. Crossovers increase genetic diversity but their main function is to ensure accurate chromosome segregation. Defects in crossover number and position produce aneuploidies that represent the main cause of miscarriages and chromosomal abnormalities such as Down's syndrome. Recombination is initiated by the formation of programmed double strand breaks (DSBs), which occur preferentially at places called DSB hotspots. Among all DSBs generated, only a small fraction is repaired by crossover, the other being repaired by other homologous recombination pathways. Crossover maps have been generated in a number of organisms, defining crossover hotspots. With the availability of genome-wide maps of DSBs as well as the ability to measure genetically the repair outcome at several hotspots, it is becoming more and more clear that not all DSB hotspots behave the same for crossover formation, suggesting that chromosomal features distinguish different types of hotspots.     

Construction of two recombination yeast two-hybrid vectors by in vitro recombination

Recombination-based restrictionless, ligation-independent cloning has been proven to be advantageous over restriction digestion and ligation cloning. To utilize the recombination cloning and previously constructed two-hybrid cDNA libraries, a new Gateway yeast two-hybrid bait vector, pEZY202, and a new prey vector, pEZY45, were constructed. The two-hybrid vectors were generated by in vitro recombination using a protocol that can be easily adapted for the conversion of other existing vectors. The new vectors were used to assay the interaction between the WW domain of PQBP1 (PQBPww) and the WW domain binding protein WBP11. Both PQBPww and WBP11 were cloned into a Gateway donor vector by in vitro recombination. They were then subcloned into pEZY45 and pEZY202, respectively, by in vitro recombination. The binding between PQBPww and WBP11 was reported in a two-hybrid experiment using the new vectors. The results of testing the new vectors in combination with the original vectors indicated that the new bait vector could be used to screen cDNA libraries that are constructed using the original prey vectors.     

NAR Breakthrough Article: Crystal structure of Hop2–Mnd1 and mechanistic insights into its role in meiotic recombination

In meiotic DNA recombination, the Hop2−Mnd1 complex promotes Dmc1-mediated single-stranded DNA (ssDNA) invasion into homologous chromosomes to form a synaptic complex by a yet-unclear mechanism. Here, the crystal structure of Hop2−Mnd1 reveals that it forms a curved rod-like structure consisting of three leucine zippers and two kinked junctions. One end of the rod is linked to two juxtaposed winged-helix domains, and the other end is capped by extra α-helices to form a helical bundle-like structure. Deletion analysis shows that the helical bundle-like structure is sufficient for interacting with the Dmc1-ssDNA nucleofilament, and molecular modeling suggests that the curved rod could be accommodated into the helical groove of the nucleofilament. Remarkably, the winged-helix domains are juxtaposed at fixed relative orientation, and their binding to DNA is likely to perturb the base pairing according to molecular simulations. These findings allow us to propose a model explaining how Hop2−Mnd1 juxtaposes Dmc1-bound ssDNA with distorted recipient double-stranded DNA and thus facilitates strand invasion.     

A cyanobacterial gene in nonphotosynthetic protists--an early chloroplast acquisition in eukaryotes?

Since the incorporation of mitochondria and chloroplasts (plastids) into the eukaryotic cell by endosymbiosis, genes have been transferred from the organellar genomes to the nucleus of the host, via an ongoing process known as endosymbiotic gene transfer. Accordingly, in photosynthetic eukaryotes, nuclear genes with cyanobacterial affinity are believed to have originated from endosymbiotic gene transfer from chloroplasts. Analysis of the Arabidopsis thaliana genome has shown that a significant fraction (2%-9%) of the nuclear genes have such an endosymbiotic origin. Recently, it was argued that 6-phosphogluconate dehydrogenase (gnd)-the second enzyme in the oxidative pentose phosphate pathway-was one such example. Here we show that gnd genes with cyanobacterial affinity also are present in several nonphotosynthetic protistan lineages, such as Heterolobosea, Apicomplexa, and parasitic Heterokonta. Current data cannot definitively resolve whether these groups acquired the gnd gene by primary and/or secondary endosymbiosis or via an independent lateral gene transfer event. Nevertheless, our data suggest that chloroplasts were introduced into eukaryotes much earlier than previously thought and that several major groups of heterotrophic eukaryotes have secondarily lost photosynthetic plastids.     

How closely does genetic diversity in finite populations conform to predictions of neutral theory? Large deficits in regions of low recombination

Levels of genetic diversity in finite populations are crucial in conservation and evolutionary biology. Genetic diversity is required for populations to evolve and its loss is related to inbreeding in random mating populations, and thus to reduced population fitness and increased extinction risk. Neutral theory is widely used to predict levels of genetic diversity. I review levels of genetic diversity in finite populations in relation to predictions of neutral theory. Positive associations between genetic diversity and population size, as predicted by neutral theory, are observed for microsatellites, allozymes, quantitative genetic variation and usually for mitochondrial DNA (mtDNA). However, there are frequently significant deviations from neutral theory owing to indirect selection at linked loci caused by balancing selection, selective sweeps and background selection. Substantially lower genetic diversity than predicted under neutrality was found for chromosomes with low recombination rates and high linkage disequilibrium (compared with 'normally' recombining chromosomes within species and adjusted for different copy numbers and mutation rates), including W (median 100% lower) and Y (89% lower) chromosomes, dot fourth chromosomes in Drosophila (94% lower) and mtDNA (67% lower). Further, microsatellite genetic and allelic diversity were lost at 12 and 33% faster rates than expected in populations adapting to captivity, owing to widespread selective sweeps. Overall, neither neutral theory nor most versions of the genetic draft hypothesis are compatible with all empirical results.