Survival differences and associated molecular signatures of DNMT3A-mutant acute myeloid leukemia patients

Acute myeloid leukemia (AML) is a very heterogeneous and highly malignant blood cancer. Mutations of the DNA methyltransferase DNMT3A are among the most frequent recurrent genetic lesions in AML. The majority of DNMT3A-mutant AML patients shows fast relapse and poor survival, but also patients with long survival or long-term remission have been reported. Underlying molecular signatures and mechanisms that contribute to these survival differences are only poorly understood and have not been studied in detail so far. We applied hierarchical clustering to somatic gene mutation profiles of 51 DNMT3A-mutant patients from The Cancer Genome Atlas (TCGA) AML cohort revealing two robust patient subgroups with profound differences in survival. We further determined molecular signatures that distinguish both subgroups. Our results suggest that FLT3 and/or NPM1 mutations contribute to survival differences of DNMT3A-mutant patients. We observed an upregulation of genes of the p53, VEGF and DNA replication pathway and a downregulation of genes of the PI3K-Akt pathway in short- compared to long-lived patients. We identified that the majority of measured miRNAs was downregulated in the short-lived group and we found differentially expressed microRNAs between both subgroups that have not been reported for AML so far (miR-153-2, miR-3065, miR-95, miR-6718) suggesting that miRNAs could be important for prognosis. In addition, we learned gene regulatory networks to predict potential major regulators and found several genes and miRNAs with known roles in AML pathogenesis, but also interesting novel candidates involved in the regulation of hematopoiesis, cell cycle, cell differentiation, and immunity that may contribute to the observed survival differences of both subgroups and could therefore be important for prognosis. Moreover, the characteristic gene mutation and expression signatures that distinguished short- from long-lived patients were also predictive for independent DNMT3A-mutant AML patients from other cohorts and could also contribute to further improve the European LeukemiaNet (ELN) prognostic scoring system. Our study represents the first in-depth computational approach to identify molecular factors associated with survival differences of DNMT3A-mutant AML patients and could trigger additional studies to develop robust molecular markers for a better stratification of AML patients with DNMT3A mutations

Database Species-Area Relationships in Palaearctic Grasslands.

The database collects the data resulting from the sampling of species-area relationships (SARs) in grassland communities in the Palaearctic. The core features of the database are the complete data from the EDGG Research Expeditions, but similar data from other studies are also included. Main features of the majority of relevés are: (i) they are part of nested-plot series (typically 0.0001-100 m²), (ii) bryophytes and lichens are treated comprehensively; (iii) detailed soil and other environmental data

The Cinderella syndrome:why do malaria-infected cells burst at midnight?

An interesting quirk of many malaria infections is that all parasites within a host-millions of them-progress through their cell cycle synchronously. This surprising coordination has long been recognized, yet there is little understanding of what controls it or why it has evolved. Interestingly, the conventional explanation for coordinated development in other parasite species does not seem to apply here. We argue that for malaria parasites, a critical question has yet to be answered: is the coordination due to parasites bursting at the same time or at a particular time? We explicitly delineate these fundamentally different scenarios, possible underlying mechanistic explanations and evolutionary drivers, and discuss the existing corroborating data and key evidence needed to solve this evolutionary mystery. © 2012 Elsevier Ltd

Generation of therapeutic antisera for emerging viral infections

The recent Ebola virus outbreak has highlighted the therapeutic potential of antisera and renewed interest in this treatment approach. While human convalescent sera may not be readily available in the early stages of an outbreak, antisera of animal origin can be produced in a short time frame. Here, we compared adjuvanted virus-like particles (VLP) with recombinant modified vaccinia virus Ankara and vesicular stomatitis virus (VSV), both expressing the Ebola virus antigens. The neutralizing antibody titers of rabbits immunized with adjuvanted VLPs were similar to those immunized with the replication-competent VSV, indicating that presentation of the antigen in its native conformation rather than de novo antigen expression is essential for production of functional antibodies. This approach also yielded high-titer antisera against Nipah virus glycoproteins, illustrating that it is transferable to other virus families. Multiple-step immunoglobulin G purification using a two-step 20-40% ammonium sulfate precipitation followed by protein A affinity chromatography resulted in 90% recovery of functionality and sustained in vivo stability. Adjuvanted VLP-based immunization strategies are thus a promising approach for the rapid generation of therapeutic antisera against emerging infections

The design and function of birds’ nests

All birds construct nests in which to lay eggs and/or raise offspring. Traditionally, it was thought that natural selection and the requirement to minimize the risk of predation determined the design of completed nests. However, it is becoming increasingly apparent that sexual selection also influences nest design. This is an important development as while species such as bowerbirds build structures that are extended phenotypic signals whose sole purpose is to attract a mate, nests contain eggs and/or offspring, thereby suggesting a direct tradeoff between the conflicting requirements of natural and sexual selection. Nest design also varies adaptively in order to both minimize the detrimental effects of parasites and to create a suitable microclimate for parents and developing offspring in relation to predictable variation in environmental conditions. Our understanding of the design and function of birds’ nests has increased considerably in recent years, and the evidence suggests that nests have four nonmutually exclusive functions. Consequently, we conclude that the design of birds’ nests is far more sophisticated than previously realized and that nests are multifunctional structures that have important fitness consequences for the builder/s

Scale-dependent plant diversity in Palaearctic grasslands: a comparative overview

Here we present an extensive overview of plant diversity values in Palaearctic grasslands for seven standard grain sizes from 0.0001 to 100 m². The data originate from 20 studies, including the Field Workshops of the Eurasian Dry Grassland Group (EDGG), ranging geographically from Spain in the west to Siberia in the east, from Sicily in the south to Estonia in the north and from the sea coast up to 3100 m a.s.l. The majority of data is from dry grasslands (Festuco-Brometea, Koelerio-Corynephoretea, Cleistogenetea squarrosae), but there are also some mesic, wet, saline, acidic, alpine and Mediterranean grasslands included. Among others, we compiled data from 1795 1-m², 1109 10-m² and 338 100-m² plots. In all cases we present mean, minimum and maximum richness for the seven grain sizes, plus, in cases where also terricolous bryophytes and lichens had been recorded, the same values for total “plant” species richness, non-vascular plant species richness and fraction of non-vascular plants. The maximum richness values were 82, 101 and 134 for all “plants”, and 79, 98 and 127 vascular plants at grain sizes of 1 m², 10 m² and 100 m², respectively (all in Transylvania, Romania). Our overview comprises new, hitherto unpublished world records of vascular plant species richness at the scales of 0.0001 m² (9) and 0.001 m² (19, both shoot presence), from meso-xeric, basiphilous grasslands in Navarre, Spain, which is much higher than the previously known maxima. The highest values of non-vascular plant richness at 1 m², 10 m² and 100 m², respectively, were 49, 64 and 64, respectively (all in Sedo-Scleranthenea communities of Öland, Sweden, and Saaremaa, Estonia). In general, the dry, alpine and Mediterranean grasslands were much richer than the studied mesic, wet or saline grasslands at any spatial scale. The presented set of mean, minimum and maximum values and their metadata is publically available and will be continuously updated. These data can serve as a reference of “normal” richness, both in fundamental and applied research. To facilitate the application, we provide an easy formula based on the power-law species-area relationship that allows the estimation of richness values at intermediate grain sizes not included in our dataset. In conclusion, our data emphasise the role of Palaearctic grasslands as global hotspot of small-scale vascular plant diversity, while at the same time highlighting that in some grassland types also the bryophyte and lichen diversity can be extraordinarily high

Proteomic profiling reveals CDK6 upregulation as a targetable resistance mechanism for lenalidomide in multiple myeloma

The immunomodulatory drugs (IMiDs) lenalidomide and pomalidomide are highly effective treatments for multiple myeloma. However, virtually all patients eventually relapse due to acquired drug resistance with resistance-causing genetic alterations being found only in a small subset of cases. To identify non-genetic mechanisms of drug resistance, we here perform integrated global quantitative tandem mass tag (TMT)-based proteomic and phosphoproteomic analyses and RNA sequencing in five paired pre-treatment and relapse samples from multiple myeloma patients. These analyses reveal a CDK6-governed protein resistance signature that includes myeloma high-risk factors such as TRIP13 and RRM1. Overexpression of CDK6 in multiple myeloma cell lines reduces sensitivity to IMiDs while CDK6 inhibition by palbociclib or CDK6 degradation by proteolysis targeting chimeras (PROTACs) is highly synergistic with IMiDs in vitro and in vivo. This work identifies CDK6 upregulation as a druggable target in IMiD-resistant multiple myeloma and highlights the use of proteomic studies to uncover non-genetic resistance mechanisms in cancer

Comprehensive CRISPR-Cas9 screens identify genetic determinants of drug responsiveness in multiple myeloma

The introduction of new drugs in the past years has substantially improved outcome in multiple myeloma (MM). However, the majority of patients eventually relapse and become resistant to one or multiple drugs. While the genetic landscape of relapsed/ resistant multiple myeloma has been elucidated, the causal relationship between relapse-specific gene mutations and the sensitivity to a given drug in MM has not systematically been evaluated. To determine the functional impact of gene mutations, we performed combined whole-exome sequencing (WES) of longitudinal patient samples with CRISPR-Cas9 drug resistance screens for lenalidomide, bortezomib, dexamethasone, and melphalan. WES of longitudinal samples from 16 MM patients identified a large number of mutations in each patient that were newly acquired or evolved from a small subclone (median 9, range 1-55), including recurrent mutations in TP53, DNAH5, and WSCD2. Focused CRISPR-Cas9 resistance screens against 170 relapse-specific mutations functionally linked 15 of them to drug resistance. These included cereblon E3 ligase complex members for lenalidomide, structural genes PCDHA5 and ANKMY2 for dexamethasone, RB1 and CDK2NC for bortezomib, and TP53 for melphalan. In contrast, inactivation of genes involved in the DNA damage repair pathway, including ATM, FANCA, RAD54B, and BRCC3, enhanced susceptibility to cytotoxic chemotherapy. Resistance patterns were highly drug specific with low overlap and highly correlated with the treatment-dependent clonal evolution in patients. The functional association of specific genetic alterations with drug sensitivity will help to personalize treatment of MM in the future

Cellular Entry of Ebola Virus Involves Uptake by a Macropinocytosis-Like Mechanism and Subsequent Trafficking through Early and Late Endosomes

Zaire ebolavirus (ZEBOV), a highly pathogenic zoonotic virus, poses serious public health, ecological and potential bioterrorism threats. Currently no specific therapy or vaccine is available. Virus entry is an attractive target for therapeutic intervention. However, current knowledge of the ZEBOV entry mechanism is limited. While it is known that ZEBOV enters cells through endocytosis, which of the cellular endocytic mechanisms used remains unclear. Previous studies have produced differing outcomes, indicating potential involvement of multiple routes but many of these studies were performed using noninfectious surrogate systems such as pseudotyped retroviral particles, which may not accurately recapitulate the entry characteristics of the morphologically distinct wild type virus. Here we used replication-competent infectious ZEBOV as well as morphologically similar virus-like particles in specific infection and entry assays to demonstrate that in HEK293T and Vero cells internalization of ZEBOV is independent of clathrin, caveolae, and dynamin. Instead the uptake mechanism has features of macropinocytosis. The binding of virus to cells appears to directly stimulate fluid phase uptake as well as localized actin polymerization. Inhibition of key regulators of macropinocytosis including Pak1 and CtBP/BARS as well as treatment with the drug EIPA, which affects macropinosome formation, resulted in significant reduction in ZEBOV entry and infection. It is also shown that following internalization, the virus enters the endolysosomal pathway and is trafficked through early and late endosomes, but the exact site of membrane fusion and nucleocapsid penetration in the cytoplasm remains unclear. This study identifies the route for ZEBOV entry and identifies the key cellular factors required for the uptake of this filamentous virus. The findings greatly expand our understanding of the ZEBOV entry mechanism that can be applied to development of new therapeutics as well as provide potential insight into the trafficking and entry mechanism of other filoviruses