Recognition of bacterial effector AvrRps4 in Arabidopsis

Pflanzen und Tiere werden ständig von Krankheitserregern, wie z.B. Viren, Bakterien und Pilzen angegriffen. Im Gegensatz zu Tieren besitzen Pflanzen kein humorales Immunsystem, sondern nur ein angeborenes Immunsystem. Krankheitserreger verfügen über Effektorproteine, welche die pflanzliche Immunantwort unterdrücken können. Um dem entgegenzuwirken, haben Pflanzen Resistenzproteine entwickelt, die entweder Effektorproteine direkt erkennen können oder deren Funktionen in der Wirtszelle. Eine bestimmte Gruppe dieser Resistenzproteine, TIR-NB-LRR Rezeptoren, benötigen das Pflanzenprotein EDS1 zur Aktivierung der Immunantwort. Aktivierung der Resistenzantwort durch die Erkennung eines Effektorproteins führt zu einem Einstrom von Kalziumionen in die Zelle, gefolgt von der lokalen Freisetzung radikaler Sauerstoffspezies und oft zur Aktivierung des programmierten Zelltods in den angegriffenen Zellen. Allerdings wird die Aktivierung des programmierten Zelltods nicht immer für die Resistenz benötigt. Um ein besseres Verständnis der Co-Evolution von Krankheitserregern und Pflanzen zu erlangen, ist es wichtig, die molekularen Mechnismen der Erkennung von Effektorproteinen sowie deren Funktionen in der Wirtszelle zu untersuchen. Desweiteren ist bisher unbekannt, wie TIR-NBLRR Rezeptoren nach ihrer Aktivierung die EDS1-abhängige Immunantwort auslösen. Diese Arbeit beschäftigt sich mit den molekularen Mechanismen der Erkennung des Effektorproteins AvrRps4 von Pseudomonas syringae durch die TIR-NB-LRR Rezeptorproteine RPS4 und RRS1 in der Wirtspflanze Arabidopsis thaliana. Eine Lokalisierung von RPS4 im Zellkern wird benötigt, um eine effektive Immunantwort auszulösen. Bisher ist nicht bekannt, in welchem Zellkompartiment die Erkennung von AvrRps4 stattfindet. Ein verstärkter Export von AvrRps4 aus dem Zellkern korreliert mit einer schwächeren Immunantwort der Pflanze wohingegen der Import von AvrRps4 in den Zellkern Resistenz auslöst. Interessanterweise führt der Import von AvrRps4 in den Zellkern zu einer vollständigen Resistenz der Pflanze, jedoch ohne das dafür programmierter Zelltod ausgelöst werden muss. Aus diesen Ergebnissen lässt sich schliessen, dass zumindest ein essentieller Schritt zur Erkennung von AvrRps4 im Zellkern stattfindet. Weitere Ergebnisse deuten darauf hin, dass die beiden Rezeptorproteine RPS4 und RRS1 kooperativ arbeiten um Resistenz gegenüber AvrRps4 zu vermitteln. Um einen besseres Verständnis für die Funktion von AvrRps4 in der Pflanze und die Erkennung durch RPS4 und RRS1 zu bekommen, wurden die Interaktionspartner von AvrRps4 untersucht. AvrRps4 interagiert mit EDS1 im Zellkern der Pflanze, was darauf hindeutet, dass EDS1 das Target (Zielprotein) von AvrRps4 sein könnte. Desweiteren wurde eine Interaktion von EDS1 mit RPS4 und RRS1 im Zellkern beobachtet. Diese Ergebnisse weisen darauf hin, dass sowohl die Erkennung von Effektorproteinen als auch die Aktivierung der Immunantwort im Zellkern der Pflanze stattfinden können

Austrian Students’ Perceptions of Social Distancing and Their Emotional Experiences During Distance Learning Due to the COVID-19 Pandemic

The COVID-19 pandemic has led to far-reaching changes in various aspects of students’ lives. In the particular case of the educational field, classroom teaching was drastically shifted to a distance learning format. Consequently, students needed to deal with a completely different everyday school life. Learning was carried out online, which implied that students could not see some of their regular, important social contacts such as teachers, classmates or friends. It is already proven that such social distancing measures had impacts on students’ emotional experiences during the lockdown. Following a mixed-methods concurrent single-phase design, this study examined students’ perceptions of social distancing and their emotional experiences during the first school lockdown in Austria. Data from an online survey (n = 263 students) and from qualitative interviews (n = 56 students) were analyzed. The results show that distance learning drastically reduced interaction among students and between students and their teachers. Furthermore, the results indicate negative emotional experiences due to less social contact, increased learning pressures, and less structure. However, findings revealed that students also experience and perceived positive emotional experiences during distance learning because of more freedom, autonomy, and to some extent, less performance pressure. Based on the findings, the present study discusses possible perspectives on how to support students during and after distance learning, as well as further lines of research.Peer Reviewe

SUMO modification of PCNA is controlled by DNA

Post-translational modification by the ubiquitin-like protein SUMO is often regulated by cellular signals that restrict the modification to appropriate situations. Nevertheless, many SUMO-specific ligases do not exhibit much target specificity, and—compared with the diversity of sumoylation substrates—their number is limited. This raises the question of how SUMO conjugation is controlled in vivo. We report here an unexpected mechanism by which sumoylation of the replication clamp protein, PCNA, from budding yeast is effectively coupled to S phase. We find that loading of PCNA onto DNA is a prerequisite for sumoylation in vivo and greatly stimulates modification in vitro. To our surprise, however, DNA binding by the ligase Siz1, responsible for PCNA sumoylation, is not strictly required. Instead, the stimulatory effect of DNA on conjugation is mainly attributable to DNA binding of PCNA itself. These findings imply a change in the properties of PCNA upon loading that enhances its capacity to be sumoylated

Cell cycle, oncogenic and tumor suppressor pathways regulate numerous long and macro non-protein-coding RNAs

Background: The genome is pervasively transcribed but most transcripts do not code for proteins, constituting non-protein-coding RNAs. Despite increasing numbers of functional reports of individual long non-coding RNAs (lncRNAs), assessing the extent of functionality among the non-coding transcriptional output of mammalian cells remains intricate. In the protein-coding world, transcripts differentially expressed in the context of processes essential for the survival of multicellular organisms have been instrumental in the discovery of functionally relevant proteins and their deregulation is frequently associated with diseases. We therefore systematically identified lncRNAs expressed differentially in response to oncologically relevant processes and cell-cycle, p53 and STAT3 pathways, using tiling arrays. Results: We found that up to 80% of the pathway-triggered transcriptional responses are non-coding. Among these we identified very large macroRNAs with pathway-specific expression patterns and demonstrated that these are likely continuous transcripts. MacroRNAs contain elements conserved in mammals and sauropsids, which in part exhibit conserved RNA secondary structure. Comparing evolutionary rates of a macroRNA to adjacent protein-coding genes suggests a local action of the transcript. Finally, in different grades of astrocytoma, a tumor disease unrelated to the initially used cell lines, macroRNAs are differentially expressed. Conclusions: It has been shown previously that the majority of expressed non-ribosomal transcripts are non-coding. We now conclude that differential expression triggered by signaling pathways gives rise to a similar abundance of non-coding content. It is thus unlikely that the prevalence of non-coding transcripts in the cell is a trivial consequence of leaky or random transcription events

Pilot Study on Mass Spectrometry–Based Analysis of the Proteome of CD34+CD123+ Progenitor Cells for the Identification of Potential Targets for Immunotherapy in Acute Myeloid Leukemia

Targeting of leukemic stem cells with specific immunotherapy would be an ideal approach for the treatment of myeloid malignancies, but suitable epitopes are unknown. The comparative proteome-level characterization of hematopoietic stem and progenitor cells from healthy stem cell donors and patients with acute myeloid leukemia has the potential to reveal differentially expressed proteins which can be used as surface-markers or as proxies for affected molecular pathways. We employed mass spectrometry methods to analyze the proteome of the cytosolic and the membrane fraction of CD34 and CD123 co-expressing FACS-sorted leukemic progenitors from five patients with acute myeloid leukemia. As a reference, CD34+CD123+ normal hematopoietic progenitor cells from five healthy, granulocyte-colony stimulating factor (G-CSF) mobilized stem cell donors were analyzed. In this Tandem Mass Tag (TMT) 10-plex labelling–based approach, 2070 proteins were identified with 171 proteins differentially abundant in one or both cellular compartments. This proof-of-principle-study demonstrates the potential of mass spectrometry to detect differentially expressed proteins in two compartment fractions of the entire proteome of leukemic stem cells, compared to their non-malignant counterparts. This may contribute to future immunotherapeutic target discoveries and individualized AML patient characterization

Salvage treatment with plerixafor in poor mobilizing allogeneic stem cell donors: results of a prospective phase II-trial

We conducted a prospective clinical trial to investigate the safety and efficacy of plerixafor (P) in allogeneic peripheral blood stem cells (PBSC) donors with poor mobilization response to standard-dose granulocyte colony-stimulating factor (G-CSF), defined by <2 × 1