Dysregulation of the high mobility group AT-hook 2 (HMGA2) gene in human tumours

The high mobility group AT-hook 2 (HMGA2) gene is expressed during embryogenesis but silenced in adult tissues. It often is reactivated for varying reasons depending on tissue type in a variety of malignant and benign tumours contributing to tumour growth. In this thesis, causes for dysregulation in various human tissues were investigated. A potential HMGA2 increase upon growth factor incubation was investigated in the epithelial prostate cancer cell line PC-3 as well as in human umbilical vein endothelial cells. The underlying mechanism for HMGA2 silencing in the prostate cancer cell line LNCaP was examined as well as effects of HMGA2 on cell viability. Furthermore, a small subset of uterine leiomyomas was tested by high-resolution array-based comparative genomic hybridisation to detect microdeletions potentially related to the breakpoints accompanying the typical translocation responsible for HMGA2 reactivation. An HMGA2 increase in PC-3 cells upon growth factor and FBS incubation was not detected, whereat it was shown in HUVECs upon growth factor incubation. HMGA2 in non-expressing LNCaP cells was found to be detectable after incubation with a demethylating agent, but not after silencing of DICER1, pointing rather to the involvement of DNA methylation than of miRNAs in HMGA2 silencing. Furthermore, HMGA2 incubation was associated with reduced cell viability. In two of the three uterine leiomyomas, small deletions that may be associated with the translocation were identified upstream of the HMGA2 locus. The results obtained herein underline the versatility of HMGA2 in regulation and function supporting the need for individual consideration for the development of potential therapeutic applications

YIP1 family member 4 (YIPF4) is a novel cellular binding partner of the papillomavirus E5 proteins

Papillomaviruses (PVs) are capable of causing a broad spectrum of diseases with the human PVs (HPVs) being responsible for a great portion of cervical, anogenital and head and neck cancers worldwide. The PV oncoprotein E5 plays roles in host cell transformation, the PV life-cycle and viral immune evasion. However, the mechanisms by which E5 achieves this are unclear. A yeast two-hybrid screen identified a novel Golgi protein, YIPF4, as a potential interactor of 16E5. YIPF4 is a member of the integral membrane protein family YIP1 that is thought to mediate intracellular trafficking. Quantitative polymerase chain reaction, Western blot and immuno-histochemistry analysis confirmed that YIPF4 is expressed in host cells of HPV infection in cell culture systems and in clinical samples of HPV16 induced cervical lesions. This implies that YIPF4 could be a relevant in vivo binding partner of E5. Upon the differentiation of HPV18 positive keratinoctyes in semisolid medium, the YIPF4 expression levels were stabilised compared to control cells suggesting that YIPF4 might play a role during the productive viral life-cycle. A differential, detergent permeabilisation assay provided the first experimental evidence for a three trans-membrane domain model of YIPF4. Co-immuno-precipitation revealed a conserved interaction of YIPF4 with E5 proteins from clinically important PVs indicating a potentially invaluable role of this complex for the virus. A flow cytometry approach unexpectedly revealed that neither E5 nor YIPF4 proteins modulate the trafficking of human leukocyte antigen class I molecules to facilitate viral immune evasion. A preliminary cellular interactome of YIPF4 was determined in a label free mass spectrometry experiment to facilitate the search for the function of the highly conserved E5/YIPF4 protein complex. This knowledge might contribute to elucidating new targets for the development of therapeutic agents against the broad spectrum of PV associated diseases

Agnoprotein Is an Essential Egress Factor during BK Polyomavirus Infection.

BK polyomavirus (BKPyV; hereafter referred to as BK) causes a lifelong chronic infection and is associated with debilitating disease in kidney transplant recipients. Despite its importance, aspects of the virus life cycle remain poorly understood. In addition to the structural proteins, the late region of the BK genome encodes for an auxiliary protein called agnoprotein. Studies on other polyomavirus agnoproteins have suggested that the protein may contribute to virion infectivity. Here, we demonstrate an essential role for agnoprotein in BK virus release. Viruses lacking agnoprotein fail to release from host cells and do not propagate to wild-type levels. Despite this, agnoprotein is not essential for virion infectivity or morphogenesis. Instead, agnoprotein expression correlates with nuclear egress of BK virions. We demonstrate that the agnoprotein binding partner α-soluble N-ethylmaleimide sensitive fusion (NSF) attachment protein (α-SNAP) is necessary for BK virion release, and siRNA knockdown of α-SNAP prevents nuclear release of wild-type BK virions. These data highlight a novel role for agnoprotein and begin to reveal the mechanism by which polyomaviruses leave an infected cell

Aspirin use and bleeding events during thrombocytopenia after autologous stem-cell transplantation for multiple myeloma

BackgroundIn patients with cardiovascular (CV) comorbidities that necessitate antiplatelet therapy (APT), its optimal management during chemotherapy-induced thrombocytopenia remains elusive, as the risk of bleeding has to be balanced against the risk of CV events. The purpose of this study was to assess the risk for bleeding with APT during thrombocytopenia in patients with multiple myeloma undergoing high-dose chemotherapy and subsequent autologous stem-cell transplantation (ASCT) with and without acetylsalicylic acid (ASA) as comedication.MethodsWe assessed patients who underwent ASCT at the Heidelberg University Hospital between 2011 and 2020 for bleeding events, management strategies for ASA intake during thrombocytopenia, transfusion requirements, and the occurrence of CV events.ResultsThere were 57/1,113 patients who continued ASA until at least 1 day after ASCT; thus, a continuous platelet inhibition during thrombocytopenia was assumed. Most of the patients (41/57) continued ASA until they had a platelet count of 20–50/nl. This range reflects the kinetics of thrombocytopenia and nondaily measurements of platelets during ASCT. A tendency toward a higher risk for bleeding events in the ASA group was demonstrated (1.9% (control group) vs. 5.3% (ASA), p = 0.082). The risk factors for bleeding in multivariate analysis were the duration of thrombocytopenia < 50/nl, a history of gastrointestinal bleeding, and diarrhea. The factors predicting the duration of thrombocytopenia were age >60 years, a hematopoietic stem-cell transplantation comorbidity index ≥3, and an impaired bone marrow reserve at admission. CV events occurred in three patients; none of them took ASA or had an indication for APT.ConclusionsThe intake of ASA until thrombocytopenia with a platelet count of 20–50/nl appears safe, although an elevated risk cannot be excluded. If ASA is indicated for the secondary prevention of CV events, the evaluation of risk factors for bleeding and a prolonged time of thrombocytopenia before conditioning is crucial to adapt the strategy for ASA intake during thrombocytopenia

ECRG4 is a candidate tumor suppressor gene frequently hypermethylated in colorectal carcinoma and glioma

<p>Abstract</p> <p>Background</p> <p>Cancer cells display widespread changes in DNA methylation that may lead to genetic instability by global hypomethylation and aberrant silencing of tumor suppressor genes by focal hypermethylation. In turn, altered DNA methylation patterns have been used to identify putative tumor suppressor genes.</p> <p>Methods</p> <p>In a methylation screening approach, we identified <it>ECRG4 </it>as a differentially methylated gene. We analyzed different cancer cells for <it>ECRG4 </it>promoter methylation by COBRA and bisulfite sequencing. Gene expression analysis was carried out by semi-quantitative RT-PCR. The <it>ECRG4 </it>coding region was cloned and transfected into colorectal carcinoma cells. Cell growth was assessed by MTT and BrdU assays. ECRG4 localization was analyzed by fluorescence microscopy and Western blotting after transfection of an <it>ECRG4-eGFP </it>fusion gene.</p> <p>Results</p> <p>We found a high frequency of <it>ECRG4 </it>promoter methylation in various cancer cell lines. Remarkably, aberrant methylation of <it>ECRG4 </it>was also found in primary human tumor tissues, including samples from colorectal carcinoma and from malignant gliomas. <it>ECRG4 </it>hypermethylation associated strongly with transcriptional silencing and its expression could be re-activated <it>in vitro </it>by demethylating treatment with 5-aza-2'-deoxycytidine. Overexpression of <it>ECRG4 </it>in colorectal carcinoma cells led to a significant decrease in cell growth. In transfected cells, ECRG4 protein was detectable within the Golgi secretion machinery as well as in the culture medium.</p> <p>Conclusions</p> <p><it>ECRG4 </it>is silenced via promoter hypermethylation in different types of human cancer cells. Its gene product may act as inhibitor of cell proliferation in colorectal carcinoma cells and may play a role as extracellular signaling molecule.</p

"Sind Sie immer noch Lehrerin?"

Das ist eine der Standardfragen von ehemaligen Schülerinnen, denen ich auf der Strasse begegne. Ich bin nach 23 Jahren immer noch als Lehrerin an der Sekundarschule tätig und übe den Beruf gerne aus. Im Folgenden beschreibe ich meine berufliche Entwicklung und frage mich, was mich in meiner Entwicklung unterstützt bzw. gefördert hat. nNach meiner Seminarausbildung in Zofingen begann mein Berufsalltag im schönen Städtchen Klingnau an der letzten Ausländerabteilung des Kantons Aargau. Ich betreute eine mehrklassige Unterstufe mit einzelnen Schülern aus Mittel- oder Oberstufe. Die Kinder sollten nach einer intensiven Einführung in einer Kleingruppe möglichst schnell den Regelklassen zugeführt werden. Mir gefiel die Arbeit mit den Schulanfängern zwar sehr, doch zugleich fühlte ich mich unterfordert. So begann ich an der Sekundarschule Englisch zu unterrichten und absolvierte gleichzeitig berufsbegleitend den Didaktikkurs an der Lehramtsschule des Kantons Aargau, um die entsprechende Qualifikation zu erwerben. Als die Integrationsklasse nach zwei Jahren aufgehoben wurde, wechselte ich an die Sekundarschule, die auch 1978 zuwenig ausgebildete Lehrkräfte aufwies. Drei Jahre lang liess man mich ohne die notwendige Französischqualifikation an der Stufe gewähren, denn Sekundarlehrkräfte blieben Mangelware. Nach einem Jahr Französischstudium in Neuchâtel kehrte ich als Sekundarlehrerin nach Klingnau zurück. Die wunderschöne Lage am Stausee, das tolle Team, die gute Infrastruktur, die ländliche Familienidylle hielten mich weitere Jahre an der Sekundarschule in Klingnau

Dysregulation des High Mobility Group AT-Hook 2 (HMGA2) Gens in humanen Tumoren

The high mobility group AT-hook 2 (HMGA2) gene is expressed during embryogenesis but silenced in adult tissues. It often is reactivated for varying reasons depending on tissue type in a variety of malignant and benign tumours contributing to tumour growth. In this thesis, causes for dysregulation in various human tissues were investigated. A potential HMGA2 increase upon growth factor incubation was investigated in the epithelial prostate cancer cell line PC-3 as well as in human umbilical vein endothelial cells. The underlying mechanism for HMGA2 silencing in the prostate cancer cell line LNCaP was examined as well as effects of HMGA2 on cell viability. Furthermore, a small subset of uterine leiomyomas was tested by high-resolution array-based comparative genomic hybridisation to detect microdeletions potentially related to the breakpoints accompanying the typical translocation responsible for HMGA2 reactivation. An HMGA2 increase in PC-3 cells upon growth factor and FBS incubation was not detected, whereat it was shown in HUVECs upon growth factor incubation. HMGA2 in non-expressing LNCaP cells was found to be detectable after incubation with a demethylating agent, but not after silencing of DICER1, pointing rather to the involvement of DNA methylation than of miRNAs in HMGA2 silencing. Furthermore, HMGA2 incubation was associated with reduced cell viability. In two of the three uterine leiomyomas, small deletions that may be associated with the translocation were identified upstream of the HMGA2 locus. The results obtained herein underline the versatility of HMGA2 in regulation and function supporting the need for individual consideration for the development of potential therapeutic applications