Nepfix: on-demand cloud elastic simulation of networks of evolutionary processors

Una Red de Procesadores Evolutivos o NEP (por sus siglas en ingles), es un modelo computacional inspirado por el modelo evolutivo de las celulas, específicamente por las reglas de multiplicación de las mismas. Esta inspiración hace que el modelo sea una abstracción sintactica de la manipulation de information de las celulas. En particu¬lar, una NEP define una maquina de cómputo teorica capaz de resolver problemas NP completos de manera eficiente en tóerminos de tiempo. En la praóctica, se espera que las NEP simuladas en móaquinas computacionales convencionales puedan resolver prob¬lemas reales complejos (que requieran ser altamente escalables) a cambio de una alta complejidad espacial. En el modelo NEP, las cóelulas estóan representadas por palabras que codifican sus secuencias de ADN. Informalmente, en cualquier momento de cómputo del sistema, su estado evolutivo se describe como un coleccion de palabras, donde cada una de ellas representa una celula. Estos momentos fijos de evolucion se denominan configuraciones. De manera similar al modelo biologico, las palabras (celulas) mutan y se dividen en base a bio-operaciones sencillas, pero solo aquellas palabras aptas (como ocurre de forma parecida en proceso de selection natural) seran conservadas para la siguiente configuracióon. Una NEP como herramienta de computation, define una arquitectura paralela y distribuida de procesamiento simbolico, en otras palabras, una red de procesadores de lenguajes. Desde el momento en que el modelo fue propuesto a la comunidad científica en el año 2001, múltiples variantes se han desarrollado y sus propiedades respecto a la completitud computacional, eficiencia y universalidad han sido ampliamente estudiadas y demostradas. En la actualidad, por tanto, podemos considerar que el modelo teórico NEP se encuentra en el estadio de la madurez. La motivación principal de este Proyecto de Fin de Grado, es proponer una aproxi-mación práctica que permita dar un salto del modelo teórico NEP a una implantación real que permita su ejecucion en plataformas computacionales de alto rendimiento, con el fin de solucionar problemas complejos que demanda la sociedad actual. Hasta el momento, las herramientas desarrolladas para la simulation del modelo NEP, si bien correctas y con resultados satisfactorios, normalmente estón atadas a su entorno de ejecucion, ya sea el uso de hardware específico o implementaciones particulares de un problema. En este contexto, el propósito fundamental de este trabajo es el desarrollo de Nepfix, una herramienta generica y extensible para la ejecucion de cualquier algo¬ritmo de un modelo NEP (o alguna de sus variantes), ya sea de forma local, como una aplicación tradicional, o distribuida utilizando los servicios de la nube. Nepfix es una aplicacion software desarrollada durante 7 meses y que actualmente se encuentra en su segunda iteration, una vez abandonada la fase de prototipo. Nepfix ha sido disenada como una aplicacion modular escrita en Java 8 y autocontenida, es decir, no requiere de un entorno de ejecucion específico (cualquier maquina virtual de Java es un contenedor vólido). Nepfix contiene dos componentes o móodulos. El primer móodulo corresponde a la ejecución de una NEP y es por lo tanto, el simulador. Para su desarrollo, se ha tenido en cuenta el estado actual del modelo, es decir, las definiciones de los procesadores y filtros mas comunes que conforman la familia del modelo NEP. Adicionalmente, este componente ofrece flexibilidad en la ejecucion, pudiendo ampliar las capacidades del simulador sin modificar Nepfix, usando para ello un lenguaje de scripting. Dentro del desarrollo de este componente, tambióen se ha definido un estóandar de representacióon del modelo NEP basado en el formato JSON y se propone una forma de representation y codificación de las palabras, necesaria para la comunicación entre servidores. Adicional-mente, una característica importante de este componente, es que se puede considerar una aplicacion aislada y por tanto, la estrategia de distribution y ejecución son total-mente independientes. El segundo moódulo, corresponde a la distribucióon de Nepfix en la nube. Este de-sarrollo es el resultado de un proceso de i+D, que tiene una componente científica considerable. Vale la pena resaltar el desarrollo de este modulo no solo por los resul-tados prócticos esperados, sino por el proceso de investigation que se se debe abordar con esta nueva perspectiva para la ejecución de sistemas de computación natural. La principal característica de las aplicaciones que se ejecutan en la nube es que son gestionadas por la plataforma y normalmente se encapsulan en un contenedor. En el caso de Nepfix, este contenedor es una aplicacion Spring que utiliza el protocolo HTTP o AMQP para comunicarse con el resto de instancias. Como valor añadido, Nepfix aborda dos perspectivas de implementation distintas (que han sido desarrolladas en dos iteraciones diferentes) del modelo de distribution y ejecucion, que tienen un impacto muy significativo en las capacidades y restricciones del simulador. En concreto, la primera iteration utiliza un modelo de ejecucion asincrono. En esta perspectiva asincrona, los componentes de la red NEP (procesadores y filtros) son considerados como elementos reactivos a la necesidad de procesar una palabra. Esta implementation es una optimization de una topologia comun en el modelo NEP que permite utilizar herramientas de la nube para lograr un escalado transparente (en lo ref¬erente al balance de carga entre procesadores) pero produce efectos no deseados como indeterminacion en el orden de los resultados o imposibilidad de distribuir eficiente-mente redes fuertemente interconectadas. Por otro lado, la segunda iteration corresponde al modelo de ejecucion sincrono. Los elementos de una red NEP siguen un ciclo inicio-computo-sincronizacion hasta que el problema se ha resuelto. Esta perspectiva sincrona representa fielmente al modelo teórico NEP pero el proceso de sincronizacion es costoso y requiere de infraestructura adicional. En concreto, se requiere un servidor de colas de mensajes RabbitMQ. Sin embargo, en esta perspectiva los beneficios para problemas suficientemente grandes superan a los inconvenientes, ya que la distribuciín es inmediata (no hay restricciones), aunque el proceso de escalado no es trivial. En definitiva, el concepto de Nepfix como marco computacional se puede considerar satisfactorio: la tecnología es viable y los primeros resultados confirman que las carac-terísticas que se buscaban originalmente se han conseguido. Muchos frentes quedan abiertos para futuras investigaciones. En este documento se proponen algunas aproxi-maciones a la solucion de los problemas identificados como la recuperacion de errores y la division dinamica de una NEP en diferentes subdominios. Por otra parte, otros prob-lemas, lejos del alcance de este proyecto, quedan abiertos a un futuro desarrollo como por ejemplo, la estandarización de la representación de las palabras y optimizaciones en la ejecucion del modelo síncrono. Finalmente, algunos resultados preliminares de este Proyecto de Fin de Grado han sido presentados recientemente en formato de artículo científico en la "International Work-Conference on Artificial Neural Networks (IWANN)-2015" y publicados en "Ad-vances in Computational Intelligence" volumen 9094 de "Lecture Notes in Computer Science" de Springer International Publishing. Lo anterior, es una confirmation de que este trabajo mas que un Proyecto de Fin de Grado, es solo el inicio de un trabajo que puede tener mayor repercusion en la comunidad científica. Abstract Network of Evolutionary Processors -NEP is a computational model inspired by the evolution of cell populations, which might model some properties of evolving cell communities at the syntactical level. NEP defines theoretical computing devices able to solve NP complete problems in an efficient manner. In this model, cells are represented by words which encode their DNA sequences. Informally, at any moment of time, the evolutionary system is described by a collection of words, where each word represents one cell. Cells belong to species and their community evolves according to mutations and division which are defined by operations on words. Only those cells are accepted as surviving (correct) ones which are represented by a word in a given set of words, called the genotype space of the species. This feature is analogous with the natural process of evolution. Formally, NEP is based on an architecture for parallel and distributed processing, in other words, a network of language processors. Since the date when NEP was pro¬posed, several extensions and variants have appeared engendering a new set of models named Networks of Bio-inspired Processors (NBP). During this time, several works have proved the computational power of NBP. Specifically, their efficiency, universality, and computational completeness have been thoroughly investigated. Therefore, we can say that the NEP model has reached its maturity. The main motivation for this End of Grade project (EOG project in short) is to propose a practical approximation that allows to close the gap between theoretical NEP model and a practical implementation in high performing computational platforms in order to solve some of high the high complexity problems society requires today. Up until now tools developed to simulate NEPs, while correct and successful, are usu¬ally tightly coupled to the execution environment, using specific software frameworks (Hadoop) or direct hardware usage (GPUs). Within this context the main purpose of this work is the development of Nepfix, a generic and extensible tool that aims to execute algorithms based on NEP model and compatible variants in a local way, similar to a traditional application or in a distributed cloud environment. Nepfix as an application was developed during a 7 month cycle and is undergoing its second iteration once the prototype period was abandoned. Nepfix is designed as a modular self-contained application written in Java 8, that is, no additional external dependencies are required and it does not rely on an specific execution environment, any JVM is a valid container. Nepfix is made of two components or modules. The first module corresponds to the NEP execution and therefore simulation. During the development the current state of the theoretical model was used as a reference including most common filters and processors. Additionally extensibility is provided by the use of Python as a scripting language to run custom logic. Along with the simulation a definition language for NEP has been defined based on JSON as well as a mechanisms to represent words and their possible manipulations. NEP simulator is isolated from distribution and as mentioned before different applications that include it as a dependency are possible, the distribution of NEPs is an example of this. The second module corresponds to executing Nepfix in the cloud. The development carried a heavy R&D process since this front was not explored by other research groups until now. It's important to point out that the development of this module is not focused on results at this point in time, instead we focus on feasibility and discovery of this new perspective to execute natural computing systems and NEPs specifically. The main properties of cloud applications is that they are managed by the platform and are encapsulated in a container. For Nepfix a Spring application becomes the container and the HTTP or AMQP protocols are used for communication with the rest of the instances. Different execution perspectives were studied, namely asynchronous and synchronous models were developed for solving different kind of problems using NEPs. Different limitations and restrictions manifest in both models and are explored in detail in the respective chapters. In conclusion we can consider that Nepfix as a computational framework is suc-cessful: Cloud technology is ready for the challenge and the first results reassure that the properties Nepfix project pursued were met. Many investigation branches are left open for future investigations. In this EOG implementation guidelines are proposed for some of them like error recovery or dynamic NEP splitting. On the other hand other interesting problems that were not in the scope of this project were identified during development like word representation standardization or NEP model optimizations. As a confirmation that the results of this work can be useful to the scientific com-munity a preliminary version of this project was published in The International Work- Conference on Artificial Neural Networks (IWANN) in May 2015. Development has not stopped since that point and while Nepfix in it's current state can not be consid¬ered a final product the most relevant ideas, possible problems and solutions that were produced during the seven months development cycle are worthy to be gathered and presented giving a meaning to this EOG work.

The folded plate model in structural analysis. Elastic, plastic and dynamic formulations for nonprismatic structures

A specific numerical procedure for the analysis of arbitrary nonprismatic folded plate structures is presented. An elastic model is studied and compared with a harmonic solution for a prismatic structure. An extension to the plastic analysis is developed, and the influence of the structural geometry and loading pattern is analyzed. Nonprismatic practical cases, with arbitrary geometry and loading are shown, as well in the elastic range as in the plastic one. Finally, a dynamic formulation is outlined

Transient excess of MYC activity can elicit genomic instability and tumorigenesis

Overexpression of the MYC protooncogene has been implicated in the genesis of diverse human tumors. Tumorigenesis induced by MYC has been attributed to sustained effects on proliferation and differentiation. Here we report that MYC may also contribute to tumorigenesis by destabilizing the cellular genome. A transient excess of MYC activity increased tumorigenicity of Rat1A cells by at least 50-fold. The increase persisted for >30 days after the return of MYC activity to normal levels. The brief surfeit of MYC activity was accompanied by evidence of genomic instability, including karyotypic abnormalities, gene amplification, and hypersensitivity to DNA-damaging agents. MYC also induced genomic destabilization in normal human fibroblasts, although these cells did not become tumorigenic. Stimulation of Rat1A cells with MYC accelerated their passage through G1/S. Moreover, MYC could force normal human fibroblasts to transit G1 and S after treatment with N-(phosphonoacetyl)-l-aspartate (PALA) at concentrations that normally lead to arrest in S phase by checkpoint mechanisms. Instead, the cells subsequently appeared to arrest in G2. We suggest that the accelerated passage through G1 was mutagenic but that the effect of MYC permitted a checkpoint response only after G2 had been reached. Thus, MYC may contribute to tumorigenesis through a dominant mutator effect.

Instability of signaling resolution models of parent–offspring conflict

Recent signaling resolution models of parent–offspring conflict have provided an important framework for theoretical and empirical studies of communication and parental care. According to these models, signaling of need is stabilized by its cost. However, our computer simulations of the evolutionary dynamics of chick begging and parental investment show that in Godfray’s model the signaling equilibrium is evolutionarily unstable: populations that start at the signaling equilibrium quickly depart from it. Furthermore, the signaling and nonsignaling equilibria are linked by a continuum of equilibria where chicks above a certain condition do not signal and we show that, contrary to intuition, fitness increases monotonically as the proportion of young that signal decreases. This result forces us to reconsider much of the current literature on signaling of need and highlights the need to investigate the evolutionary stability of signaling equilibria based on the handicap principle.

Induction of microsatellite instability by oxidative DNA damage

Instability of repetitive sequences, both in intronic sequences and within coding regions, has been demonstrated to be a hallmark of genomic instability in human cancer. Understanding how these mutational events arise may provide an opportunity for prevention or early intervention in cancer development. To study the source of this instability, we have identified a region of the β-lactamase gene that is tolerant to the insertion of fragments of exogenous DNA as large as 1,614 bp with minimal loss of enzyme activity, as determined by antibiotic resistance. Fragments inserted out-of-frame render Escherichia coli sensitive to antibiotic, and compensatory frameshift mutations that restore the reading frame of β-lactamase can be selected on the basis of antibiotic resistance. We have utilized this site to insert a synthetic microsatellite sequence within the β-lactamase gene and selected for mutations yielding frameshifts. This assay provides for detection of one frameshift mutation in a background of 106 wild-type sequences. Mismatch repair deficiency increased the observed frameshift frequency ≈300-fold. Exposure of plasmid containing microsatellite sequences to hydrogen peroxide resulted in frameshift mutations that were localized exclusively to the microsatellite sequences, whereas DNA damage by UV or N-methyl-N′-nitro-N-nitrosoguanidine did not result in enhanced mutagenesis. We postulate that in tumor cells, endogenous production of oxygen free radicals may be a major factor in promoting instability of microsatellite sequences. This β-lactamase assay may provide a sensitive methodology for the detection and quantitation of mutations associated with the development of cancer.

The human XRCC9 gene corrects chromosomal instability and mutagen sensitivities in CHO UV40 cells

The Chinese hamster ovary (CHO) mutant UV40 cell line is hypersensitive to UV and ionizing radiation, simple alkylating agents, and DNA cross-linking agents. The mutant cells also have a high level of spontaneous chromosomal aberrations and 3-fold elevated sister chromatid exchange. We cloned and sequenced a human cDNA, designated XRCC9, that partially corrected the hypersensitivity of UV40 to mitomycin C, cisplatin, ethyl methanesulfonate, UV, and γ-radiation. The spontaneous chromosomal aberrations in XRCC9 cDNA transformants were almost fully corrected whereas sister chromatid exchanges were unchanged. The XRCC9 genomic sequence was cloned and mapped to chromosome 9p13. The translated XRCC9 sequence of 622 amino acids has no similarity with known proteins. The 2.5-kb XRCC9 mRNA seen in the parental cells was undetectable in UV40 cells. The mRNA levels in testis were up to 10-fold higher compared with other human tissues and up to 100-fold higher compared with other baboon tissues. XRCC9 is a candidate tumor suppressor gene that might operate in a postreplication repair or a cell cycle checkpoint function.

Minisatellite instability in severe combined immunodeficiency mouse cells

We have recently found that okadaic acid, which shows strong inhibitory activity on protein serine/threonine phosphatases and tumor-promoting activity in vivo and in vitro, induces minisatellite mutation (MSM). Human tumors and chemically induced counterparts in experimental animals are also sometimes associated with MSM. In the present study, we demonstrated minisatellite (MS) instability in severe combined immunodeficiency (SCID) cells in which the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is impaired. Cells from a SCID fibroblast cell line transformed by simian virus 40 large tumor antigen, SC3VA2, and from an embryonal SCID fibroblast cell line, SC1K, were cloned and propagated to 107 to 108 cells, and then subjected to subcloning. After propagation of each subclone to 107 to 108 cells, DNA samples were digested with HinfI and analyzed by Southern blotting using the Pc-1 MS sequence as a probe. Under low-stringency conditions, about 40 MS bands were detected, with 45% ± 6% and 37% ± 3% of SC3VA2 and SC1K cells, respectively, having MSM. In contrast, cells from the RD13B2 cell line, which was established from SCVA2 by introducing human chromosome 8q fragments, on which DNA-PKcs is known to reside, to complement the SCID phenotype, showed a very low frequency of MSM (3% ± 3%). The high frequencies of MSM in SC3VA2 and SC1K were significant, with no difference between the two. The present study clearly demonstrates that MS instability exists in SCID fibroblasts, suggesting that DNA-PKcs might be involved in the stable maintenance of MS sequences in the genome.

The onset and extent of genomic instability in sporadic colorectal tumor progression

Cancer cell genomes contain alterations beyond known etiologic events, but their total number has been unknown at even the order of magnitude level. By sampling colorectal premalignant polyp and carcinoma cell genomes through use of the technique inter-(simple sequence repeat) PCR, we have found genomic alterations to be considerably more abundant than expected, with the mean number of genomic events per carcinoma cell totaling approximately 11,000. Colonic polyps early in the tumor progression pathway showed similar numbers of events. These results indicate that, as with certain hereditary cancer syndromes, genomic destabilization is an early step in sporadic tumor development. Together these results support the model of genomic instability being a cause rather than an effect of malignancy, facilitating vastly accelerated somatic cell evolution, with the observed orderly steps of the colon cancer progression pathway reflecting the consequences of natural selection.

Genetic instability of cancer cells is proportional to their degree of aneuploidy

Genetic and phenotypic instability are hallmarks of cancer cells, but their cause is not clear. The leading hypothesis suggests that a poorly defined gene mutation generates genetic instability and that some of many subsequent mutations then cause cancer. Here we investigate the hypothesis that genetic instability of cancer cells is caused by aneuploidy, an abnormal balance of chromosomes. Because symmetrical segregation of chromosomes depends on exactly two copies of mitosis genes, aneuploidy involving chromosomes with mitosis genes will destabilize the karyotype. The hypothesis predicts that the degree of genetic instability should be proportional to the degree of aneuploidy. Thus it should be difficult, if not impossible, to maintain the particular karyotype of a highly aneuploid cancer cell on clonal propagation. This prediction was confirmed with clonal cultures of chemically transformed, aneuploid Chinese hamster embryo cells. It was found that the higher the ploidy factor of a clone, the more unstable was its karyotype. The ploidy factor is the quotient of the modal chromosome number divided by the normal number of the species. Transformed Chinese hamster embryo cells with a ploidy factor of 1.7 were estimated to change their karyotype at a rate of about 3% per generation, compared with 1.8% for cells with a ploidy factor of 0.95. Because the background noise of karyotyping is relatively high, the cells with low ploidy factor may be more stable than our method suggests. The karyotype instability of human colon cancer cell lines, recently analyzed by Lengnauer et al. [Lengnauer, C., Kinzler, K. W. & Vogelstein, B. (1997) Nature (London) 386, 623–627], also corresponds exactly to their degree of aneuploidy. We conclude that aneuploidy is sufficient to explain genetic instability and the resulting karyotypic and phenotypic heterogeneity of cancer cells, independent of gene mutation. Because aneuploidy has also been proposed to cause cancer, our hypothesis offers a common, unique mechanism of altering and simultaneously destabilizing normal cellular phenotypes.

Evolutionary instability of the major histocompatibility complex class I loci in New World primates

Homologues of the human major histocompatibility complex (MHC) HLA-A, -B, -E, -F, and -G loci are present in all the Catarrhini (Old World primates, apes, and humans), and some of their allelic lineages have survived several speciation events. Analysis of 26 MHC class I cDNAs from seven different genera of New World primates revealed that the Callitrichinae (tamarins and marmosets) are an exception to these rules of MHC stability. In gene trees of primate MHC class I genes, sequences from the Callitrichinae cluster in a genus-specific fashion, whereas in the other genera of New World primates, as in the Catarrhini, they cluster in a transgeneric way. The genus-specific clustering of the Callitrichinae cDNAs indicates that there is no orthology between MHC class I loci in genera of this phyletic group. Additionally, the Callitrichinae genera exhibit limited variability of their MHC class I genes, in contrast to the high variability displayed by all other primates. Each Callitrichinae genus, therefore, expresses its own set of MHC class I genes, suggesting that an unusually high rate of turnover of loci occurs in this subfamily. The limited variability of MHC class I genes in the Callitrichinae is likely the result of the recent origin of these loci.

The DNA Helicase Activity of BLM Is Necessary for the Correction of the Genomic Instability of Bloom Syndrome Cells

Bloom syndrome (BS) is a rare autosomal recessive disorder characterized by growth deficiency, immunodeficiency, genomic instability, and the early development of cancers of many types. BLM, the protein encoded by BLM, the gene mutated in BS, is localized in nuclear foci and absent from BS cells. BLM encodes a DNA helicase, and proteins from three missense alleles lack displacement activity. BLM transfected into BS cells reduces the frequency of sister chromatid exchanges and restores BLM in the nucleus. Missense alleles fail to reduce the sister chromatid exchanges in transfected BS cells or restore the normal nuclear pattern. BLM complements a phenotype of a Saccharomyces cerevisiae sgs1 top3 strain, and the missense alleles do not. This work demonstrates the importance of the enzymatic activity of BLM for its function and nuclear localization pattern.

Characterization of an In Vitro Model of Elastic Fiber Assembly

Elastic fibers consist of two morphologically distinct components: elastin and 10-nm fibrillin-containing microfibrils. During development, the microfibrils form bundles that appear to act as a scaffold for the deposition, orientation, and assembly of tropoelastin monomers into an insoluble elastic fiber. Although microfibrils can assemble independent of elastin, tropoelastin monomers do not assemble without the presence of microfibrils. In the present study, immortalized ciliary body pigmented epithelial (PE) cells were investigated for their potential to serve as a cell culture model for elastic fiber assembly. Northern analysis showed that the PE cells express microfibril proteins but do not express tropoelastin. Immunofluorescence staining and electron microscopy confirmed that the microfibril proteins produced by the PE cells assemble into intact microfibrils. When the PE cells were transfected with a mammalian expression vector containing a bovine tropoelastin cDNA, the cells were found to express and secrete tropoelastin. Immunofluorescence and electron microscopic examination of the transfected PE cells showed the presence of elastic fibers in the matrix. Biochemical analysis of this matrix showed the presence of cross-links that are unique to mature insoluble elastin. Together, these results indicate that the PE cells provide a unique, stable in vitro system in which to study elastic fiber assembly.

Chromosomal instability and cytoskeletal defects in oral cancer cells

Oral squamous cell carcinomas are characterized by complex, often near-triploid karyotypes with structural and numerical variations superimposed on the initial clonal chromosomal alterations. We used immunohistochemistry combined with classical cytogenetic analysis and spectral karyotyping to investigate the chromosomal segregation defects in cultured oral squamous cell carcinoma cells. During division, these cells frequently exhibit lagging chromosomes at both metaphase and anaphase, suggesting defects in the mitotic apparatus or kinetochore. Dicentric anaphase chromatin bridges and structurally altered chromosomes with consistent long arms and variable short arms, as well as the presence of gene amplification, suggested the occurrence of breakage–fusion–bridge cycles. Some anaphase bridges were observed to persist into telophase, resulting in chromosomal exclusion from the reforming nucleus and micronucleus formation. Multipolar spindles were found to various degrees in the oral squamous cell carcinoma lines. In the multipolar spindles, the poles demonstrated different levels of chromosomal capture and alignment, indicating functional differences between the poles. Some spindle poles showed premature splitting of centrosomal material, a precursor to full separation of the microtubule organizing centers. These results indicate that some of the chromosomal instability observed within these cancer cells might be the result of cytoskeletal defects and breakage–fusion–bridge cycles.

The human papillomavirus type 16 E6 and E7 oncoproteins cooperate to induce mitotic defects and genomic instability by uncoupling centrosome duplication from the cell division cycle

Loss of genomic integrity is a defining feature of many human malignancies, including human papillomavirus (HPV)-associated preinvasive and invasive genital squamous lesions. Here we show that aberrant mitotic spindle pole formation caused by abnormal centrosome numbers represents an important mechanism in accounting for numeric chromosomal alterations in HPV-associated carcinogenesis. Similar to what we found in histopathological specimens, HPV-16 E6 and E7 oncoproteins cooperate to induce abnormal centrosome numbers, aberrant mitotic spindle pole formation, and genomic instability. The low-risk HPV-6 E6 and E7 proteins did not induce such abnormalities. Whereas the HPV-16 E6 oncoprotein has no immediate effects on centrosome numbers, HPV-16 E7 rapidly induces abnormal centrosome duplication. Thus our results suggest a model whereby HPV-16 E7 induces centrosome-related mitotic disturbances that are potentiated by HPV-16 E6.

Separating Growth from Elastic Deformation during Cell Enlargement1

Plants change size by deforming reversibly (elastically) whenever turgor pressure changes, and by growing. The elastic deformation is independent of growth because it occurs in nongrowing cells. Its occurrence with growth has prevented growth from being observed alone. We investigated whether the two processes could be separated in internode cells of Chara corallina Klien ex Willd., em R.D.W. by injecting or removing cell solution with a pressure probe to change turgor while the cell length was continuously measured. Cell size changed immediately when turgor changed, and growth rates appeared to be altered. Low temperature eliminated growth but did not alter the elastic effects. This allowed elastic deformation measured at low temperature to be subtracted from elongation at warm temperature in the same cell. After the subtraction, growth alone could be observed for the first time. Alterations in turgor caused growth to change rapidly to a new, steady rate with no evidence of rapid adjustments in wall properties. This turgor response, together with the marked sensitivity of growth to temperature, suggested that the growth rate was not controlled by inert polymer extension but rather by biochemical reactions that include a turgor-sensitive step.