Embedded Linux integration using Linuxlink tool and peripheral driver development for Zedboard

This project is divided into two main parts: The first part shows the integration of an Embedded Linux operating system on a development hardware platform named Zedboard. This platform contains a Zynq-7000 System on Chip (Soc) which is composed by two dual core ARM Cortex-A9 processors and a FPGA Artix-7. The Embedded Linux is built with Linuxlink, a Timesys tool. Meanwhile, the platform hardware configuration is done with Xilinx Vivado. The system is loaded with an SD card which requires to have every files needed for the booting process and for the operation. Some of these files are generated with Xilinx SDK software. The second part starts up from the system already built to integrate a peripheral in the Zynq-7000 FPGA. Also the drivers for controlling the peripheral from the operating system are developed. Finally, a user space program is created to test both of them. RESUMEN. Este proyecto consta de dos partes: La primera muestra la integración de un sistema operativo Linux embebido en una plataforma de desarrollo hardware llamada Zedboard. Esta plataforma utiliza un System on Chip (SoC) Zynq-7000 que está formado por dos procesadores ARM Cortex-A9 de doble núcleo y una FPGA Artix-7. El Linux embebido se construye utilizando la herramienta Linuxlink de Timesys, mientras que el hardware de la plataforma de desarrollo se configura con Vivado de Xilinx. El sistema se carga en una tarjeta SD que debe tener todos los archivos necesarios para completar el arranque y hacer funcionar el sistema. Algunos de esos archivos se generan con la herramienta SDK de Xilinx. En la segunda parte se utiliza el sistema construido para integrar un periférico en la FPGA del Zynq-7000, haciendo uso de Vivado, y se desarrollan los drivers necesarios para utilizarlo mediante el sistema operativo. Para probar esta última parte se desarrolla un programa de espacio de usuario.

Study of OLSR protocol with Network Simulator 2 over Linux

Until a few years ago, most of the network communications were based in the wire as the physical media, but due to the advances and the maturity of the wireless communications, this is changing. Nowadays wireless communications offers fast, secure, efficient and reliable connections. Mobile communications are in expansion, clearly driven by the use of smart phones and other mobile devices, the use of laptops, etc… Besides that point, the inversion in the installation and maintenance of the physical medium is much lower than in wired communications, not only because the air has no cost, but because the installation and maintenance of the wire require a high economic cost. Besides the economic cost we find that wire is a more vulnerable medium to external threats such as noise, sabotages, etc… There are two different types of wireless networks: those which the structure is part of the network itself and those which have a lack of structure or any centralization, in a way that the devices that form part of the network can connect themselves in a dynamic and random way, handling also the routing of every control and information messages, this kind of networks is known as Ad-hoc. In the present work we will proceed to study one of the multiple wireless protocols that allows mobile communications, it is Optimized Link State Routing, from now on, OLSR, it is an pro-active routing, standard mechanism that works in a distributed in order to stablish the connections among the different nodes that belong to a wireless network. Thanks to this protocol it is possible to get all the routing tables in all the devices correctly updated every moment through the periodical transmission of control messages and on this way allow a complete connectivity among the devices that are part of the network and also, allow access to other external networks such as virtual private networks o Internet. This protocol could be perfectly used in environments such as airports, malls, etc… The update of the routing tables in all the devices is got thanks to the periodical transmission of control messages and finally it will offer connectivity among all the devices and the corresponding external networks. For the study of OLSR protocol we will have the help of the network simulator “Network Simulator 2”, a freeware network simulator programmed in C++ based in discrete events. This simulator is used mainly in educational and research environments and allows a very extensive range of protocols, both, wired networks protocols and wireless network protocols, what is going to be really useful to proceed to the simulation of different configurations of networks and protocols. In the present work we will also study different simulations with Network Simulator 2, in different scenarios with different configurations, wired networks, and Ad-hoc networks, where we will study OLSR Protocol. RESUMEN. Hasta hace pocos años, la mayoría de las comunicaciones de red estaban basadas en el cable como medio físico pero debido al avance y madurez alcanzados en el campo de las comunicaciones inalámbricas esto está cambiando. Hoy día las comunicaciones inalámbricas nos ofrecen conexiones veloces, seguras, eficientes y fiables. Las comunicaciones móviles se encuentran en su momento de máxima expansión, claramente impulsadas por el uso de teléfonos y demás dispositivos móviles, el uso de portátiles, etc… Además la inversión a realizar en la instalación y el mantenimiento del medio físico en las comunicaciones móviles es muchísimo menor que en comunicaciones por cable, ya no sólo porque el aire no tenga coste alguno, sino porque la instalación y mantenimiento del cable precisan de un elevado coste económico por norma. Además del coste económico nos encontramos con que es un medio más vulnerable a amenazas externas tales como el ruido, escuchas no autorizadas, sabotajes, etc… Existen dos tipos de redes inalámbricas: las constituidas por una infraestructura que forma parte más o menos de la misma y las que carecen de estructura o centralización alguna, de modo que los dispositivos que forman parte de ella pueden conectarse de manera dinámica y arbitraria entre ellos, encargándose además del encaminamiento de todos los mensajes de control e información, a este tipo de redes se las conoce como redes Ad-hoc. En el presente Proyecto de Fin de Carrera se procederá al estudio de uno de los múltiples protocolos inalámbricos que permiten comunicaciones móviles, se trata del protocolo inalámbrico Optimized Link State Routing, de ahora en adelante OLSR, un mecanismo estándar de enrutamiento pro-activo, que trabaja de manera distribuida para establecer las conexiones entre los nodos que formen parte de las redes inalámbricas Ad-hoc, las cuales carecen de un nodo central y de una infraestructura pre-existente. Gracias a este protocolo es posible conseguir que todos los equipos mantengan en todo momento las tablas de ruta actualizadas correctamente mediante la transmisión periódica de mensajes de control y así permitir una completa conectividad entre todos los equipos que formen parte de la red y, a su vez, también permitir el acceso a otras redes externas tales como redes privadas virtuales o Internet. Este protocolo sería usado en entornos tales como aeropuertos La actualización de las tablas de enrutamiento de todos los equipos se conseguirá mediante la transmisión periódica de mensajes de control y así finalmente se podrá permitir conectividad entre todos los equipos y con las correspondientes redes externas. Para el estudio del protocolo OLSR contaremos con el simulador de redes Network Simulator 2, un simulador de redes freeware programado en C++ basado en eventos discretos. Este simulador es usado principalmente en ambientes educativos y de investigación y permite la simulación tanto de protocolos unicast como multicast. El campo donde más se utiliza es precisamente en el de la investigación de redes móviles Ad-hoc. El simulador Network Simulator 2 no sólo implementa el protocolo OLSR, sino que éste implementa una amplia gama de protocolos, tanto de redes cableadas como de redes inalámbricas, lo cual va a sernos de gran utilidad para proceder a la simulación de distintas configuraciones de redes y protocolos. En el presente Proyecto de Fin de Carrera se estudiarán también diversas simulaciones con el simulador NS2 en diferentes escenarios con diversas configuraciones; redes cableadas, redes inalámbricas Ad-hoc, donde se estudiará el protocolo antes mencionado: OLSR. Este Proyecto de Fin de Carrera consta de cuatro apartados distintos: Primeramente se realizará el estudio completo del protocolo OLSR, se verán los beneficios y contrapartidas que ofrece este protocolo inalámbrico. También se verán los distintos tipos de mensajes existentes en este protocolo y unos pequeños ejemplos del funcionamiento del protocolo OLSR. Seguidamente se hará una pequeña introducción al simulador de redes Network Simulator 2, veremos la historia de este simulador, y también se hará referencia a la herramienta extra NAM, la cual nos permitirá visualizar el intercambio de paquetes que se produce entre los diferentes dispositivos de nuestras simulaciones de forma intuitiva y amigable. Se hará mención a la plataforma MASIMUM, encargada de facilitar en un entorno académico software y documentación a sus alumnos con el fin de facilitarles la investigación y la simulación de redes y sensores Ad-hoc. Finalmente se verán dos ejemplos, uno en el que se realizará una simulación entre dos PCs en un entorno Ethernet y otro ejemplo en el que se realizará una simulación inalámbrica entre cinco dispositivos móviles mediante el protocolo a estudiar, OLSR.

GADD45 induction of a G2/M cell cycle checkpoint

G1/S and G2/M cell cycle checkpoints maintain genomic stability in eukaryotes in response to genotoxic stress. We report here both genetic and functional evidence of a Gadd45-mediated G2/M checkpoint in human and murine cells. Increased expression of Gadd45 via microinjection of an expression vector into primary human fibroblasts arrests the cells at the G2/M boundary with a phenotype of MPM2 immunopositivity, 4n DNA content and, in 15% of the cells, centrosome separation. The Gadd45-mediated G2/M arrest depends on wild-type p53, because no arrest was observed either in p53-null Li–Fraumeni fibroblasts or in normal fibroblasts coexpressed with p53 mutants. Increased expression of cyclin B1 and Cdc25C inhibited the Gadd45-mediated G2/M arrest in human fibroblasts, indicating that the mechanism of Gadd45-mediated G2/M checkpoint is at least in part through modulation of the activity of the G2-specific kinase, cyclin B1/p34cdc2. Genetic and physiological evidence of a Gadd45-mediated G2/M checkpoint was obtained by using GADD45-deficient human or murine cells. Human cells with endogenous Gadd45 expression reduced by antisense GADD45 expression have an impaired G2/M checkpoint after exposure to either ultraviolet radiation or methyl methanesulfonate but are still able to undergo G2 arrest after ionizing radiation. Lymphocytes from gadd45-knockout mice (gadd45 −/−) also retained a G2/M checkpoint initiated by ionizing radiation and failed to arrest at G2/M after exposure to ultraviolet radiation. Therefore, the mammalian genome is protected by a multiplicity of G2/M checkpoints in response to specific types of DNA damage.

DRC1, DNA replication and checkpoint protein 1, functions with DPB11 to control DNA replication and the S-phase checkpoint in Saccharomyces cerevisiae

In addition to DNA polymerase complexes, DNA replication requires the coordinate action of a series of proteins, including regulators Cdc28/Clb and Dbf4/Cdc7 kinases, Orcs, Mcms, Cdc6, Cdc45, and Dpb11. Of these, Dpb11, an essential BRCT repeat protein, has remained particularly enigmatic. The Schizosaccharomyces pombe homolog of DPB11, cut5, has been implicated in the DNA replication checkpoint as has the POL2 gene with which DPB11 genetically interacts. Here we describe a gene, DRC1, isolated as a dosage suppressor of dpb11–1. DRC1 is an essential cell cycle-regulated gene required for DNA replication. We show that both Dpb11 and Drc1 are required for the S-phase checkpoint, including the proper activation of the Rad53 kinase in response to DNA damage and replication blocks. Dpb11 is the second BRCT-repeat protein shown to control Rad53 function, possibly indicating a general function for this class of proteins. DRC1 and DPB11 show synthetic lethality and reciprocal dosage suppression. The Drc1 and Dpb11 proteins physically associate and function together to coordinate DNA replication and the cell cycle.

Protein kinase mutants of human ATR increase sensitivity to UV and ionizing radiation and abrogate cell cycle checkpoint control

In fission yeast, the rad3 gene product plays a critical role in sensing DNA structure defects and activating damage response pathways. A structural homologue of rad3 in humans (ATR) has been identified based on sequence similarity in the protein kinase domain. General information regarding ATR expression, protein kinase activity, and cellular localization is known, but its function in human cells remains undetermined. In the current study, the ATR protein was examined by gel filtration of protein extracts and was found to exist predominantly as part of a large protein complex. A kinase-inactivated form of the ATR gene was prepared by site-directed mutagenesis and was used in transfection experiments to probe the function of this complex. Introduction of this kinase-dead ATR into a normal fibroblast cell line, an ATM-deficient fibroblast line derived from a patient with ataxia–telangiectasia, or a p53 mutant cell line all resulted in significant losses in cell viability. Clones expressing the kinase-dead ATR displayed increased sensitivity to x-rays and UV and a loss of checkpoint control. We conclude that ATR functions as a critical part of a protein complex that mediates responses to ionizing and UV radiation in human cells. These responses include effects on cell viability and cell cycle checkpoint control.

Abrogation of the G2 cell cycle checkpoint associated with overexpression of HSIX1: A possible mechanism of breast carcinogenesis

While conducting a search for cell cycle-regulated genes in human mammary carcinoma cells, we identified HSIX1, a recently discovered member of a new homeobox gene subfamily. HSIX1 expression was absent at the onset of and increased toward the end of S phase. Since its expression pattern is suggestive of a role after S phase, we investigated the effect of HSIX1 in the G2 cell cycle checkpoint. Overexpression of HSIX1 in MCF7 cells abrogated the G2 cell cycle checkpoint in response to x-ray irradiation. HSIX1 expression was absent or very low in normal mammary tissue, but was high in 44% of primary breast cancers and 90% of metastatic lesions. In addition, HSIX1 was expressed in a variety of cancer cell lines, suggesting an important function in multiple tumor types. These data support the role for homeobox genes in tumorigenesis/tumor progression, possibly through a cell cycle function.

A p53-dependent S-phase checkpoint helps to protect cells from DNA damage in response to starvation for pyrimidine nucleotides

Normal mammalian cells arrest primarily in G1 in response to N-(phosphonacetyl)-l-aspartate (PALA), which starves them for pyrimidine nucleotides, and do not generate or tolerate amplification of the CAD gene, which confers resistance to PALA. Loss of p53, accompanied by loss of G1 arrest, permits CAD gene amplification and the consequent formation of PALA-resistant colonies. We have found rat and human cell lines that retain wild-type p53 but have lost the ability to arrest in G1 in response to PALA. However, these cells still fail to give PALA-resistant colonies and are protected from DNA damage through the operation of a second checkpoint that arrests them reversibly within S-phase. This S-phase arrest, unmasked in the absence of the G1 checkpoint, is dependent on p53 and independent of p21/waf1.

Mitosis in vertebrate somatic cells with two spindles: Implications for the metaphase/anaphase transition checkpoint and cleavage

During mitosis an inhibitory activity associated with unattached kinetochores prevents PtK1 cells from entering anaphase until all kinetochores become attached to the spindle. To gain a better understanding of how unattached kinetochores block the metaphase/anaphase transition we followed mitosis in PtK1 cells containing two independent spindles in a common cytoplasm. We found that unattached kinetochores on one spindle did not block anaphase onset in a neighboring mature metaphase spindle 20 μm away that lacked unattached kinetochores. As in cells containing a single spindle, anaphase onset occurred in the mature spindles x̄ = 24 min after the last kinetochore attached regardless of whether the adjacent immature spindle contained one or more unattached kinetochores. These findings reveal that the inhibitory activity associated with an unattached kinetochore is functionally limited to the vicinity of the spindle containing the unattached kinetochore. We also found that once a mature spindle entered anaphase the neighboring spindle also entered anaphase x̄ = 9 min later regardless of whether it contained monooriented chromosomes. Thus, anaphase onset in the mature spindle catalyzes a “start anaphase” reaction that spreads globally throughout the cytoplasm and overrides the inhibitory signal produced by unattached kinetochores in an adjacent spindle. Finally, we found that cleavage furrows often formed between the two independent spindles. This reveals that the presence of chromosomes and/or a spindle between two centrosomes is not a prerequisite for cleavage in vertebrate somatic cells.

The anaphase inhibitor of Saccharomyces cerevisiae Pds1p is a target of the DNA damage checkpoint pathway

Inhibition of DNA replication and physical DNA damage induce checkpoint responses that arrest cell cycle progression at two different stages. In Saccharomyces cerevisiae, the execution of both checkpoint responses requires the Mec1 and Rad53 proteins. This observation led to the suggestion that these checkpoint responses are mediated through a common signal transduction pathway. However, because the checkpoint-induced arrests occur at different cell cycle stages, the downstream effectors mediating these arrests are likely to be distinct. We have previously shown that the S. cerevisiae protein Pds1p is an anaphase inhibitor and is essential for cell cycle arrest in mitosis in the presence DNA damage. Herein we show that DNA damage, but not inhibition of DNA replication, induces the phosphorylation of Pds1p. Analyses of Pds1p phosphorylation in different checkpoint mutants reveal that in the presence of DNA damage, Pds1p is phosphorylated in a Mec1p- and Rad9p-dependent but Rad53p-independent manner. Our data place Pds1p and Rad53p on parallel branches of the DNA damage checkpoint pathway. We suggest that Pds1p is a downstream target of the DNA damage checkpoint pathway and that it is involved in implementing the DNA damage checkpoint arrest specifically in mitosis.

Schizosaccharomyces pombe cdc20+ encodes DNA polymerase ɛ and is required for chromosomal replication but not for the S phase checkpoint

In fission yeast both DNA polymerase alpha (pol α) and delta (pol δ) are required for DNA chromosomal replication. Here we demonstrate that Schizosaccharomyces pombe cdc20+ encodes the catalytic subunit of DNA polymerase epsilon (pol ɛ) and that this enzyme is also required for DNA replication. Following a shift to the restrictive temperature, cdc20 temperature-sensitive mutant cells block at the onset of DNA replication, suggesting that cdc20+ is required early in S phase very near to the initiation step. In the budding yeast Saccharomyces cerevisiae, it has been reported that in addition to its proposed role in chromosomal replication, DNA pol ɛ (encoded by POL2) also functions directly as an S phase checkpoint sensor [Navas, T. A., Zhou, Z. & Elledge, S. J. (1995) Cell 80, 29–39]. We have investigated whether cdc20+ is required for the checkpoint control operating in fission yeast, and our data indicate that pol ɛ does not have a role as a checkpoint sensor coordinating S phase with mitosis. In contrast, germinating spores disrupted for the gene encoding pol α rapidly enter mitosis in the absence of DNA synthesis, suggesting that in the absence of pol α, normal coordination between S phase and mitosis is lost. We propose that the checkpoint signal operating in S phase depends on assembly of the replication initiation complex, and that this signal is generated prior to the elongation stage of DNA synthesis.

Cdc25 Inhibited In Vivo and In Vitro by Checkpoint Kinases Cds1 and Chk1

In the fission yeast Schizosaccharomyces pombe, the protein kinase Cds1 is activated by the S–M replication checkpoint that prevents mitosis when DNA is incompletely replicated. Cds1 is proposed to regulate Wee1 and Mik1, two tyrosine kinases that inhibit the mitotic kinase Cdc2. Here, we present evidence from in vivo and in vitro studies, which indicates that Cds1 also inhibits Cdc25, the phosphatase that activates Cdc2. In an in vivo assay that measures the rate at which Cdc25 catalyzes mitosis, Cds1 contributed to a mitotic delay imposed by the S–M replication checkpoint. Cds1 also inhibited Cdc25-dependent activation of Cdc2 in vitro. Chk1, a protein kinase that is required for the G2–M damage checkpoint that prevents mitosis while DNA is being repaired, also inhibited Cdc25 in the in vitro assay. In vitro, Cds1 and Chk1 phosphorylated Cdc25 predominantly on serine-99. The Cdc25 alanine-99 mutation partially impaired the S–M replication and G2–M damage checkpoints in vivo. Thus, Cds1 and Chk1 seem to act in different checkpoint responses to regulate Cdc25 by similar mechanisms.

Regulation of Telomere Length by Checkpoint Genes in Schizosaccharomyces pombe

We have studied telomere length in Schizosaccharomyces pombe strains carrying mutations affecting cell cycle checkpoints, DNA repair, and regulation of the Cdc2 protein kinase. Telomere shortening was found in rad1, rad3, rad17, and rad26 mutants. Telomere lengths in previously characterized rad1 mutants paralleled the replication checkpoint proficiency of those mutants. In contrast, rad9, chk1, hus1, and cds1 mutants had intact telomeres. No difference in telomere length was seen in mutants affected in the regulation of Cdc2, whereas some of the DNA repair mutants examined had slightly longer telomeres than did the wild type. Overexpression of the rad1+ gene caused telomeres to elongate slightly. The kinetics of telomere shortening was monitored by following telomere length after disruption of the rad1+ gene; the rate was ∼1 nucleotide per generation. Wild-type telomere length could be restored by reintroduction of the wild-type rad1+ gene. Expression of the Saccharomyces cerevisiae RCK1 protein kinase gene, which suppresses the radiation and hydroxyurea sensitivity of Sz. pombe checkpoint mutants, was able to attenuate telomere shortening in rad1 mutant cells and to increase telomere length in a wild-type background. The functional effects of telomere shortening in rad1 mutants were assayed by measuring loss of a linear and a circular minichromosome. A minor increase in loss rate was seen with the linear minichromosome, and an even smaller difference compared with wild-type was detected with the circular plasmid.

The Human G2 Checkpoint Control Protein hRAD9 Is a Nuclear Phosphoprotein That Forms Complexes with hRAD1 and hHUS1

Eukaryotic cells actively block entry into mitosis in the presence of DNA damage or incompletely replicated DNA. This response is mediated by signal transduction cascades called cell cycle checkpoints. We show here that the human checkpoint control protein hRAD9 physically associates with two other checkpoint control proteins, hRAD1 and hHUS1. Furthermore, hRAD1 and hHUS1 themselves interact, analogously to their fission yeast homologues Rad1 and Hus1. We also show that hRAD9 is present in multiple phosphorylation forms in vivo. These phosphorylated forms are present in tissue culture cells that have not been exposed to exogenous sources of DNA damage, but it remains possible that endogenous damage or naturally occurring replication intermediates cause the observed phosphorylation. Finally, we show that hRAD9 is a nuclear protein, indicating that in this signal transduction pathway, hRAD9 is physically proximal to the upstream (DNA damage) signal rather than to the downstream, cytoplasmic, cell cycle machinery.

Effects of Genome Position and the DNA Damage Checkpoint on the Structure and Frequency of sod2 Gene Amplification in Fission Yeast

The Schizosaccharomyces pombe sod2 gene, located near the telomere on the long arm of chromosome I, encodes a Na+ (or Li+)/H+ antiporter. Amplification of sod2 has previously been shown to confer resistance to LiCl. We analyzed 20 independent LiCl-resistant strains and found that the only observed mechanism of resistance is amplification of sod2. The amplicons are linear, extrachromosomal elements either 225 or 180 kb long, containing both sod2 and telomere sequences. To determine whether proximity to a telomere is necessary for sod2 amplification, a strain was constructed in which the gene was moved to the middle of the same chromosomal arm. Selection of LiCl-resistant strains in this genetic background also yielded amplifications of sod2, but in this case the amplified DNA was exclusively chromosomal. Thus, proximity to a telomere is not a prerequisite for gene amplification in S. pombe but does affect the mechanism. Relative to wild-type cells, mutants with defects in the DNA damage aspect of the rad checkpoint control pathway had an increased frequency of sod2 amplification, whereas mutants defective in the S-phase completion checkpoint did not. Two models for generating the amplified DNA are presented.

The Spindle Checkpoint of Budding Yeast Depends on a Tight Complex between the Mad1 and Mad2 Proteins

The spindle checkpoint arrests the cell cycle at metaphase in the presence of defects in the mitotic spindle or in the attachment of chromosomes to the spindle. When spindle assembly is disrupted, the budding yeast mad and bub mutants fail to arrest and rapidly lose viability. We have cloned the MAD2 gene, which encodes a protein of 196 amino acids that remains at a constant level during the cell cycle. Gel filtration and co-immunoprecipitation analyses reveal that Mad2p tightly associates with another spindle checkpoint component, Mad1p. This association is independent of cell cycle stage and the presence or absence of other known checkpoint proteins. In addition, Mad2p binds to all of the different phosphorylated isoforms of Mad1p that can be resolved on SDS-PAGE. Deletion and mutational analysis of both proteins indicate that association of Mad2p with Mad1p is critical for checkpoint function and for hyperphosphorylation of Mad1p.