Konservierte und divergente Aspekte der twist-, snail- und concertina-Funktion im Käfer Tribolium castaneum

Ziel dieser Arbeit war eine Ausweitung des Verständnisses evolutionärer Veränderungen der Mesodermentwicklung und der Gastrulation und der damit verbundenden DV-Musterbildung bei Insekten. Zu diesem Zweck wurden die Funktionen der Gene twi, sna und concertina beim Käfer Tribolium castaneum (Tc) analysiert und mit den bekannten Funktionen aus der Fliege Drosophila melanogaster (Dm) verglichen. Bei Drosophila aktiviert der stabile Dorsal-Kern-Gradient twi und sna auf der Ventralseite des frühen Embryos. Zellen dieser Domäne invaginieren durch die Ventralfurche und bilden die mesodermale Schicht im Embryo. Andere DV-Musterbildungsgene werden ebenfalls von Dm-Dorsal in spezifischen Domänen entlang der DV-Achse des Embryos aktiviert. Bei Tribolium castaneum, einem Käfer mit einer anzestraleren Art der Embryogenese, verschwindet der Tc-Dorsal-Kern-Gradient früher als bei Drosophila, er ist aber dennoch in der Lage, Tc-twi und weitere DV-Musterbildungsgene ventral zu aktivieren. In dieser Arbeit konnte gezeigt werden, dass das zygotische Gen Tc-twi an der Terminierung des Dorsal-Gradienten beteiligt ist und somit eine wichtige Funktion beim Übergang der maternalen zur zygotischen DV-Musterbildungskontrolle spielt. Zusammen mit Tc-sna ist Tc-twi für die Etablierung der korrekten Musterbildung von Mesoderm, Mesektoderm und Neuroektoderm während der Gastrulation notwendig. Tc-Twi ist, wie auch bei Drosophila, der Hauptregulator für die Entwicklung des Mesoderms und ein Initiator von Zellmorphologieänderungen für die Ventralfurchenbildung. Des Weiteren ist Tc-sna für die Etablierung von Gewebegrenzen und die Kontrolle der Zellproliferation nötig, scheint aber keine fundamentale Rolle während der Mesodermentwicklung zu haben. Auf der Suche nach Homologen bekannter Effektorgene, die für Zellmorphologieänderungen in der Ventralfurche von Drosophila verantwortlich sind, wurde Tc-concertina (cta) identifiziert. Die essentielle Funktion von Cta bei Tribolium scheint allerdings die Bildung der posterioren Amnionfalte zu sein, durch welche die Amnionhöhle ensteht.. Ohne die Bewegung des posterioren Amnions ähnelt ein Querschnitt von gastrulierenden Tribolium Embryonen auffallend dem von Drosophila Embryonen. Die sehr ähnliche Morphologie der posterioreren Mitteldarminvagination bei Drosophila und der posterioren Amnionfaltenbildung bei Tribolium, sowie die Tatsache, dass beide Vorgänge Cta benötigen, deutet darauf hin, dass es sich um homologe morphologische Prozesse handelt

Modeling Initiation of Ewing Sarcoma in Human Neural Crest Cells

Ewing sarcoma family tumors (ESFT) are aggressive bone and soft tissue tumors that express EWS-ETS fusion genes as driver mutations. Although the histogenesis of ESFT is controversial, mesenchymal (MSC) and/or neural crest (NCSC) stem cells have been implicated as cells of origin. For the current study we evaluated the consequences of EWS-FLI1 expression in human embryonic stem cell-derived NCSC (hNCSC). Ectopic expression of EWS-FLI1 in undifferentiated hNCSC and their neuro-mesenchymal stem cell (hNC-MSC) progeny was readily tolerated and led to altered expression of both well established as well as novel EWS-FLI1 target genes. Importantly, whole genome expression profiling studies revealed that the molecular signature of established ESFT is more similar to hNCSC than any other normal tissue, including MSC, indicating that maintenance or reactivation of the NCSC program is a feature of ESFT pathogenesis. Consistent with this hypothesis, EWS-FLI1 induced hNCSC genes as well as the polycomb proteins BMI-1 and EZH2 in hNC-MSC. In addition, up-regulation of BMI-1 was associated with avoidance of cellular senescence and reversible silencing of p16. Together these studies confirm that, unlike terminally differentiated cells but consistent with bone marrow-derived MSC, NCSC tolerate expression of EWS-FLI1 and ectopic expression of the oncogene initiates transition to an ESFT-like state. In addition, to our knowledge this is the first demonstration that EWS-FLI1-mediated induction of BMI-1 and epigenetic silencing of p16 might be critical early initiating events in ESFT tumorigenesis

Self-Regulatory Circuits in Dorsoventral Axis Formation of the Short-Germ Beetle Tribolium castaneum

SummaryThe rel/NF-κB transcription factor Dorsal controls dorsoventral (DV) axis formation in Drosophila. A stable nuclear gradient of Dorsal directly regulates ∼50 target genes. In Tribolium castaneum (Tc), a beetle with an ancestral type of embryogenesis, the Dorsal nuclear gradient is not stable, but rapidly shrinks and disappears. We find that negative feedback accounts for this dynamic behavior: Tc-Dorsal and one of its target genes activate transcription of the IkB homolog Tc-cactus, terminating Dorsal function. Despite its transient role, Tc-Dorsal is strictly required to initiate DV polarity, as in Drosophila. However, unlike in Drosophila, embryos lacking Tc-Dorsal display a periodic pattern of DV cell fates along the AP axis, indicating that a self-organizing ectodermal patterning system operates independently of mesoderm or maternal DV polarity cues. Our results also elucidate how extraembryonic tissues are organized in short-germ embryos, and how patterning information is transmitted from the early embryo to the growth zone

Developmental Cell Article Self-Regulatory Circuits in Dorsoventral Axis Formation of the Short-Germ Beetle Tribolium castaneum

The rel/NF-kB transcription factor Dorsal controls dorsoventral (DV) axis formation in Drosophila. A stable nuclear gradient of Dorsal directly regulates 50 target genes. In Tribolium castaneum (Tc), a beetle with an ancestral type of embryogenesis, the Dorsal nuclear gradient is not stable, but rapidly shrinks and disappears. We find that negative feedback accounts for this dynamic behavior: Tc-Dorsal and one of its target genes activate transcription of the IkB homolog Tc-cactus, terminating Dorsal function. Despite its transient role, Tc-Dorsal is strictly required to initiate DV polarity, as in Drosophila. However, unlike in Drosophila, embryos lacking Tc-Dorsal display a periodic pattern of DV cell fates along the AP axis, indicating that a self-organizing ectodermal patterning system operates independently of mesoderm or maternal DV polarity cues. Our results also elucidate how extraembryonic tissues are organized in short-germ embryos, and how patterning information is transmitted from the early embryo to the growth zone

Fog signaling has diverse roles in epithelial morphogenesis in insects

The Drosophila Fog pathway represents one of the best-understood signaling cascades controlling epithelial morphogenesis. During gastrulation, Fog induces apical cell constrictions that drive the invagination of mesoderm and posterior gut primordia. The cellular mechanisms underlying primordia internalization vary greatly among insects and recent work has suggested that Fog signaling is specific to the fast mode of gastrulation found in some flies. On the contrary, here we show in the beetle Tribolium, whose development is broadly representative for insects, that Fog has multiple morphogenetic functions. It modulates mesoderm internalization and controls a massive posterior infolding involved in gut and extraembryonic development. In addition, Fog signaling affects blastoderm cellularization, primordial germ cell positioning, and cuboidal-to-squamous cell shape transitions in the extraembryonic serosa. Comparative analyses with two other distantly related insect species reveals that Fog's role during cellularization is widely conserved and therefore might represent the ancestral function of the pathway

Overexpression of HOX genes is prevalent in Ewing sarcoma and is associated with altered epigenetic regulation of developmental transcription programs

<div><p>The polycomb proteins BMI-1 and EZH2 are highly overexpressed by Ewing sarcoma (ES), a tumor of stem cell origin that is driven by EWS-ETS fusion oncogenes, most commonly EWS-FLI1. In the current study we analyzed expression of transcription programs that are controlled by polycomb proteins during embryonic development to determine if they are abnormal in ES. Our results show that polycomb target gene expression in ES deviates from normal tissues and stem cells and that, as expected, most targets are relatively repressed. However, we also discovered a paradoxical up regulation of numerous polycomb targets and these were highly enriched for homeobox (HOX) genes. Comparison of HOX profiles between malignant and non-malignant tissues revealed a distinctive HOX profile in ES, which was characterized by overexpression of posterior HOXD genes. In addition, ectopic expression of EWS-FLI1 during stem cell differentiation led to aberrant up regulation of posterior HOXD genes. Mechanistically, this up regulation was associated with altered epigenetic regulation. Specifically, ES and EWS-FLI1+ stem cells displayed a relative loss of polycomb-dependent H3K27me3 and gain of trithorax-dependent H3K4me3 at the promoters of posterior HOXD genes and also at the HOXD11.12 polycomb response element. In addition, a striking correlation was evident between <i>HOXD13</i> and other genes whose regulation is coordinately regulated during embryonic development by distal enhancer elements. Together, these studies demonstrate that epigenetic regulation of polycomb target genes, in particular HOXD genes, is altered in ES and that these changes are mediated downstream of EWS-FLI1.</p></div

Co-occurrence of targetable mutations in Non-small cell lung cancer (NSCLC) patients harboring MAP2K1 mutations

Background: MAP2K1 mutations are rare in non-small cell lung cancer (NSCLC) and considered to be mutually exclusive from known driver mutations. Activation of the MEK1-cascade is considered pivotal in resistance to targeted therapy approaches, and MAP2K1 K57 N mutation could be linked to resistance in preclinical models. We set out this study to detect MAP2K1 mutations and potentially targetable co-mutations using a molecular multiplex approach. Methods: Between 2012 and 2018, we routinely analyzed 14.512 NSCLC patients with two next-generation sequencing (NGS) panels. In a subset of patients, fluorescence in-situ hybridization was performed to detect rearrangements or amplifications. We assessed clinical parameters and co-occurring mutations and compared treatment outcomes of different forms of systemic therapy. Results: We identified 66 (0.5%) patients with MAP2K1 mutations. Both adenocarcinoma (n = 62) and squamous cell carcinoma (n = 4) histology. The presence of the mutations was linked to smoking, and transversions were more common than transitions. K57 N was the most frequent MAP2K1 mutation (n = 25). Additional mutations were found in 57 patients (86.4%). Mutations of TP53 were detected in 33 patients, followed by KEAP1 mutations in 28.1%. 24 patients (36.4%) had either MAP2K1-only or a co-occurring aberration considered targetable, including EGFR mutations, a BRAF V600E mutation and ROS1 rearrangements. Outcome analyses revealed a trend toward benefit from pemetrexed treatment. Conclusion: Our analysis shows that MAP2K1-mutated NSCLC patients might frequently present with potentially targetable aberrations. Their role in providing resistance in these subtypes and the possible therapeutic opportunities justify further analyses of this rare NSCLC subgroup

Cancer care in German centers of excellence during the first 2 years of the COVID-19 pandemic

Purpose An increasing number of international studies demonstrate serious negative effects of the COVID-19 pandemic on the timely diagnosis of cancer and on cancer treatment. Our study aimed to quantitatively and qualitatively evaluate the capacities of German Comprehensive Cancer Centers (CCCs) in different areas of complex oncology care during the first 2 years of the COVID-19 pandemic. Methods Prospective panel survey over 23 rounds among 18 CCCs in Germany between March 2020 and June 2022. Results The COVID-19 pandemic substantially affected the oncological care system in Germany during the first 2 years. Persistent limitations of care in CCCs primarily affected follow-up (- 21%) and psycho-oncologic care (- 12%), but also tumor surgery (- 9%). Substantial limitations were also reported for all other areas of multidisciplinary oncological care. Conclusions This study documents the limitations of oncological care during the COVID-19 pandemic and highlights the need to develop strategies to avoid similar limitations in the future