Russell-like bodies in plant seeds share common features with prolamin bodies and occur upon recombinant protein production

Although many recombinant proteins have been produced in seeds at high yields without adverse effects on the plant, endoplasmic reticulum (ER) stress and aberrant localization of endogenous or recombinant proteins have also been reported. The production of murine interleukin-10 (mIL-10) in Arabidopsis thaliana seeds resulted in the de novo formation of ER-derived structures containing a large fraction of the recombinant protein in an insoluble form. These bodies containing mIL-10 were morphologically similar to Russell bodies found in mammalian cells. We confirmed that the compartment containing mIL-10 was enclosed by ER membranes, and 3D electron microscopy revealed that these structures have a spheroidal shape. Another feature shared with Russell bodies is the continued viability of the cells that generate these organelles. To investigate similarities in the formation of Russell-like bodies and the plant-specific protein bodies formed by prolamins in cereal seeds, we crossed plants containing ectopic ER-derived prolamin protein bodies with a line accumulating mIL-10 in Russell-like bodies. This resulted in seeds containing only one population of protein bodies in which mIL-10 inclusions formed a central core surrounded by the prolamin-containing matrix, suggesting that both types of protein aggregates are together removed from the secretory pathway by a common mechanism. We propose that, like mammalian cells, plant cells are able to form Russell-like bodies as a self-protection mechanism, when they are overloaded with a partially transport-incompetent protein, and we discuss the resulting challenges for recombinant protein production

Potential utilization of blood leeches in modern veterinary medicine, a critical review

Blood leeches are natural medical products. They are used since several thousand years for treatment of very different clinical diseases. Over the years effects observed in the field determined through experience can be explained by those components that researchers were able to isolate. At the same time, there are reported effects of leech treatment that have not been confirmed by field studies so far. This thesis focuses on the analyses of the current situation of medical research and medical treatment in the field utilizing leeches. As a basis a general overview of biology of leeches is provided and legislative aspects of leeches as a medical product are defined. The main focus of this thesis is on the pharmacologic aspects of leech therapy. The most important substances that have been identified in leech saliva and their indications in human and veterinary medicine are reviewed. Furthermore, potential areas of utilizing leeches in veterinary medicine are reviewed and evaluated, with a focus on treating tendon injuries in horses

Genetic and molecular analysis of trichome patterning in natural accessions of Arabidopsis thaliana

An pflanzlichen Epidermen entwickeln sich Zellen häufig zu Trichomen (Blatthaare), ihnen werden unterschiedliche Schutzfunktionen vor biotischen und abiotischen Einflüssen zugeschrieben. A. thaliana Trichome sind ein intensiv beforschtes Modellsystem der Zellspezifikation und der Musterbildung. Anwesenheit eines spezifischen WD40/bHLH/R2R3MYB Aktivator Komplexes führt zur Trichomentwicklung, Mitglieder der single-repeat R3 MYB Proteinfamilie dagegen inhibieren die Trichomentwicklung benachbarter Zellen und wirken nicht-zellautonom. Die hier vorliegende Arbeit nutzte nun die vorhandene natürliche Variation der Trichomdichte in A. thaliana Populationen, um weitere Informationen zum molekularen Mechanismus der Trichomentwicklung zu bekommen. Eine F2 Kartierungs-Population, basierend auf zwei A. thaliana Ökotypen mit extrem unterschiedlicher Trichomdichte, wurde einer „quantitative trait locus“ (QTL) Analyse unterzogen. Unter den vier detektierten QTL Regionen befindet sich eine, die die Trichomdichte sehr stark beeinflusst und die im weiteren auf einen 8.4 cM Bereich eingeengt werden konnte. Anschließende funktionelle und populationsgenetische Experimente mit mehreren Kandidatengenen wiesen nach, dass sich das kausale „quantitative trait“ Nukleotid (QTN) in der kodierenden Region des single-repeat R3 MYB Gens ENHANCER OF TRIPTYCHON AND CAPRICE2 (ETC2) befindet. Dieses QTN führt zu einem Aminosäureaustausch eines konservierten Lysins zu Glutamat und stellt damit den ersten Hinweis auf eine mögliche Rolle des Prozesses der Ubiquitinierung in der Trichommusterbildung dar. Um ein tieferes Verständnis für den Mechanismus der differentiellen Trichominitiation zu erhalten, wurden Analysen auf zellulärer Ebene durchgeführt, die zeigten, dass die Modulation mit der Anzahl dazwischen liegender Mesophyllzellen gekoppelt ist. Dies ist ein Phänotyp, der eine Mutation in caprice (cpc), einem single-repeat R3 MYB Gen, widerspiegelt. Weiters beschreibt diese Arbeit ein neues Pilotprojekt, das einen Zusammenhang zwischen Trichomdichte, Anthocyangehalt und UV-B Toleranz in natürlichen A. thaliana Populationen untersucht, da ein WD40/bHLH/R2R3MYB Transkriptionskomplex auch Enzyme der Anthocyanbiosynthese reguliert.Trichomes (leaf hairs), epidermal cells protruding from aerial surfaces of plants, are thought to influence many aspects of plant physiology and ecology and thus may be of selective importance. The A. thaliana trichome serves as an intensely studied model system for cell specification. The current mechanistic concept explains de novo trichome patterning by local activation mediated by a specific WD40/bHLH/R2R3MYB activator complex which is counteracted by single-repeat R3 MYB protein non-cell autonomous acting. Existing natural variation for trichome density in A. thaliana provides the opportunity to complement well-established information about the genetic pathway of trichome initiation with knowledge about the modulation of trichome density in natural accessions. In this spirit, a quantitative trait locus (QTL) analysis of an F2 mapping population derived from a low and a high trichome density accession was carried out and detected four and seven QTL affecting aspects of trichome patterning and leaf morphology, respectively. Among the former a major QTL for trichome density was fine mapped to an 8.4 cM interval on chromosome two. Further population genetic and functional tests lead to the isolation of a quantitative trait nucleotide (QTN) located in ENHANCER OF TRIPTYCHON AND CAPRICE2 (ETC2), a member of the single-repeat R3 MYB gene family of trichome repressors. The QTN causes an exchange of a conserved lysine to glutamate in low trichome density accessions and therefore presents the first indication that ubiquitination may play a role in modulating trichome density. Furthermore, to better understand the mechanism of differential initiation of leaf trichomes on a mechanistic level, cellular analysis of fully grown leaves showed that a main difference between the low and the high trichome density accession is expressed by the number of cells formed between trichomes and that this phenocopies mutants in caprice (cpc), a single-repeat R3 MYB gene. Given that a WD40/bHLH/R2R3MYB transcription activator complex is also being involved in transcriptional regulation of the anthocyanin biosynthesis pathway, the second part of this work describes a pilot study investigating possible relationships between trichome density, anthocyanin content and UV-B tolerance in A. thaliana accessions.eingereicht von Julia HilscherAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersZsfassung in dt. SpracheWien, Univ. für Bodenkultur, Diss., 2011OeBB(VLID)193068

Symplectic difference systems: oscillation and spectral theory

This monograph is devoted to covering the main results in the qualitative theory of symplectic difference systems, including linear Hamiltonian difference systems and Sturm-Liouville difference equations, with the emphasis on the oscillation and spectral theory. As a pioneer monograph in this field it contains nowadays standard theory of symplectic systems, as well as the most current results in this field, which are based on the recently developed central object - the comparative index. The book contains numerous results and citations, which were till now scattered only in journal papers. The book also provides new applications of the theory of matrices in this field, in particular of the Moore-Penrose pseudoinverse matrices, orthogonal projectors, and symplectic matrix factorizations. Thus it brings this topic to the attention of researchers and students in pure as well as applied mathematics

Use of Fluorescent Protein Tags to Study Nuclear Organization of the Spliceosomal Machinery in Transiently Transformed Living Plant Cells

Although early studies suggested that little compartmentalization exists within the nucleus, more recent studies on metazoan systems have identified a still increasing number of specific subnuclear compartments. Some of these compartments are dynamic structures; indeed, protein and RNA-protein components can cycle between different domains. This is particularly evident for RNA processing components. In plants, lack of tools has hampered studies on nuclear compartmentalization and dynamics of RNA processing components. Here, we show that transient expression of fluorescent protein fusions of U1 and U2 small nuclear ribonucleoprotein particle (snRNP)-specific proteins U1-70K, U2B″, and U2A ′, nucleolar proteins Nop10 and PRH75, and serine-arginine-rich proteins in plant protoplasts results in their correct localization. Furthermore, snRNP-specific proteins also were correctly assembled into mature snRNPs. This system allowed a systematic analysis of the cellular localization of Arabidopsis serine-arginine-rich proteins, which, like their animal counterparts, localize to speckles but not to nucleoli and Cajal bodies. Finally, markers for three different nuclear compartments, namely, nucleoli, Cajal bodies, and speckles, have been established and were shown to be applicable for colocalization studies in living plant protoplasts. Thus, transient expression of proteins tagged with four different fluorescent proteins is a suitable system for studying the nuclear organization of spliceosomal proteins in living plant cells and should therefore allow studies of their dynamics as well

A preliminary study on methylphenidate-regulated gene expression in lymphoblastoid cells of ADHD patients

OBJECTIVES: Methylphenidate (MPH) is a commonly used stimulant medication for treating attention-deficit/hyperactivity disorder (ADHD). Besides inhibiting monoamine reuptake there is evidence that MPH also influences gene expression directly. METHODS: We investigated the impact of MPH treatment on gene expression levels of lymphoblastoid cells derived from adult ADHD patients and healthy controls by hypothesis-free, genome-wide microarray analysis. Significant findings were subsequently confirmed by quantitative Real-Time PCR (qRT PCR) analysis. RESULTS: The microarray analysis from pooled samples after correction for multiple testing revealed 138 genes to be marginally significantly regulated due to MPH treatment, and one gene due to diagnosis. By qRT PCR we could confirm that GUCY1B3 expression was differential due to diagnosis. We verified chronic MPH treatment effects on the expression of ATXN1, HEY1, MAP3K8 and GLUT3 in controls as well as acute treatment effects on the expression of NAV2 and ATXN1 specifically in ADHD patients. CONCLUSIONS: Our preliminary results demonstrate MPH treatment differences in ADHD patients and healthy controls in a peripheral primary cell model. Our results need to be replicated in larger samples and also using patient-derived neuronal cell models to validate the contribution of those genes to the pathophysiology of ADHD and mode of action of MPH