Dissection of the C-terminal region of E1A redefines the roles of CtBP and other cellular targets in oncogenic transformation.

Human adenovirus E1A makes extensive connections with the cellular protein interaction network. By doing so, E1A can manipulate many cellular programs, including cell cycle progression. Through these reprogramming events, E1A functions as a growth-promoting oncogene and has been used extensively to investigate mechanisms contributing to oncogenesis. Nevertheless, it remains unclear how the C-terminal region of E1A contributes to oncogenic transformation. Although this region is required for transformation in cooperation with E1B, it paradoxically suppresses transformation in cooperation with activated Ras. Previous analysis has suggested that the interaction of E1A with CtBP plays a pivotal role in both activities. However, some C-terminal mutants of E1A retain CtBP binding and yet exhibit defects in transformation, suggesting that other targets of this region are also necessary. To explore the roles of these additional factors, we performed an extensive mutational analysis of the C terminus of E1A. We identified key residues that are specifically required for binding all known targets of the C terminus of E1A. We further tested each mutant for the ability to both localize to the nucleus and transform primary rat cells in cooperation with E1B-55K or Ras. Interaction of E1A with importin α3/Qip1, dual-specificity tyrosine-regulated kinase 1A (DYRK1A), HAN11, and CtBP influenced transformation with E1B-55K. Interestingly, the interaction of E1A with DYRK1A and HAN11 appeared to play a role in suppression of transformation by activated Ras whereas interaction with CtBP was not necessary. This unexpected result suggests a need for revision of current models and provides new insight into transformation by the C terminus of E1A.

Current radioactive fallout contamination along a trans-European gradient assessed using terricolous lichens.

Lichens are considered to be good indicators of contamination of the terrestrial environment. In this study, we investigated the level of 137Cs and 4 K accumulated by Cladonia arbuscula and Stereocaulon alpinum along a longitudinal gradient from northern Norway, across Sweden to southern Poland. Additionally, we compared isotope contents between the selected lichen species, and investigated the correlation of the 137Cs content accumulated by C. arbuscula with 137Cs fallout after the Chernobyl disaster. The activity of 137Cs varied from 3.58 Bq kg-1 to 559 Bq kg-1 for S. alpinum, and from 1.18 Bq kg-1 to 130 Bq kg-1 for C. arbuscula. The activity of 4 K ranged from 114 Bq kg-1 to 341 Bq kg-1 for S. alpinum and from 27.2 Bq kg-1 to 314 Bq kg-1 for C. arbuscula. The 137Cs content did not differ between C. arbuscula and S. alpinum; however, the difference between species was significant for 4 K accumulation. The activity of 137Cs in C. arbuscula was significantly correlated with deposition from 1986. Based on our findings we created a spatial map of 137Cs activity in lichens measured 30 years after the event that was the primary source of this isotope. We showed that C. arbuscula can be used to assess contamination and create interpolation maps of radionuclide deposition, even if the primary deposition took place many years ago.

Impact of distance from the glacier on the content of 137Cs and 90Sr in the lichen Cetrariella delisei.

The Arctic region is substantially a pristine area, but this unique part of the globe has also been contaminated by anthropogenic radioactive nuclides, and now there is still measurable activity of anthropogenic isotopes, even though more than 50 years have passed since the main source. Radionuclides in the Arctic, especially 90Sr have seldom been studied despite their considerable environmental importance. This manuscript covers the results of 90Sr and 137Cs measurements in soil and lichen Cetrariella delisei collected from the Svalbard in 2012. In both lichen thalli and surface soils high activities of 90Sr and 137Cs were recorded and ranged between 3.69 and 28.1 Bq kg-1 90Sr and 5.38-280.1 Bq kg-1137Cs in thalli and between 4.53 and 12.78 Bq kg-1dw 90Sr and 60.6-426.1 Bq kg-1dw 137Cs in surface soil layer. The activity of 90Sr and 137Cs in lichen thalli was influenced by distance from the glacier. This showed that during radionuclide biomonitoring of particular area with the use of lichens, it is important to take into account influence of environmental variability on radionuclides contents.

New tomographic atom probe at University of Muenster, Germany.

Currently atom probe tomography provides the highest spatial resolution compared to all other volume analysis techniques. Owing to its single atom sensitivity, it is specially suited to study nano-structured materials. Therefore, a new atom probe was installed at the Institute for Material Physics at University of Muenster, Germany, to study thin film reactions. Since the available budget was rather limited, a cost-effective non-commercial atom probe was constructed. The instrument is based on a 2D delay line detector system of 120 mm diameter. To achieve a large collecting angle and thus large volumes of analysis, a straight flight tube without a reflectron is used. This way, the flight distance may be reduced down to 160 mm. However, the variable chamber layout allows using a reflectron as an alternative. Furthermore, a laser system is implemented that delivers pulses in the 500 ps range to make possible laser-assisted evaporation of atoms. The article describes instrumental details and presents first characteristic data.

Cloning, genomic organization and expression pattern of a novel Drosophila gene, the disco-interacting protein 2 (dip2), and its murine homolog.

We report the cloning and initial characterization of a novel gene encoding the Disco interacting protein 2 (Dip2). dip2 DNA complementary to RNA (cDNA) showed a high degree of sequence similarity to cDNAs of unknown function previously identified in humans and Caenorhabditis elegans. We have cloned the mouse homolog of the dip2 cDNA and characterized the expression of this gene by Northern blotting analysis and in situ hybridization to whole mount embryos. Our observations demonstrate that there is a remarkable degree of sequence conservation at the dip2 locus that is reflected in the nervous system-specific expression of both the Drosophila and mouse homologs.

Identification of a molecular recognition feature in the E1A oncoprotein that binds the SUMO conjugase UBC9 and likely interferes with polySUMOylation.

Hub proteins have central roles in regulating cellular processes. By targeting a single cellular hub, a viral oncogene may gain control over an entire module in the cellular interaction network that is potentially comprised of hundreds of proteins. The adenovirus E1A oncoprotein is a viral hub that interacts with many cellular hub proteins by short linear motifs/molecular recognition features (MoRFs). These interactions transform the architecture of the cellular protein interaction network and virtually reprogram the cell. To identify additional MoRFs within E1A, we screened portions of E1A for their ability to activate yeast pseudohyphal growth or differentiation. This identified a novel functional region within E1A conserved region 2 comprised of the sequence EVIDLT. This MoRF is necessary and sufficient to bind the N-terminal region of the SUMO conjugase UBC9, which also interacts with SUMO noncovalently and is involved in polySUMOylation. Our results suggest that E1A interferes with polySUMOylation, but not with monoSUMOylation. These data provide the first insight into the consequences of the interaction of E1A with UBC9, which was initially described in 1996. We further demonstrate that polySUMOylation regulates pseudohyphal growth and promyelocytic leukemia body reorganization by E1A. In conclusion, the interaction of the E1A oncogene with UBC9 mimics the normal binding between SUMO and UBC9 and represents a novel mechanism to modulate polySUMOylation.

Autoregulation of the Drosophila disconnected gene in the developing visual system.

The Drosophila disconnected (disco) gene is required for the formation of appropriate connections between the larval optic nerve and its target cells in the brain. The disco gene encodes a nuclear protein with two zinc fingers, which suggests that the gene product is a transcription factor. Here, we present data supporting this notion. We find that disco expression in the optic lobe primordium, a group of cells contacted by the developing optic nerve, depends on an autoregulatory feedback loop. We show that wild-type disco function is required for maintenance of disco mRNA and protein expression in the developing optic lobe. In addition, we demonstrate that ubiquitous Disco activity supplied by a heat-inducible gene construct activates expression from the endogenous disco gene specifically in the optic lobe primordium. Consistent with a role of Disco as a transcriptional regulatory protein, we show that portions of the Disco protein are capable of activating the transcription of reporter constructs in a heterologous system. Moreover, we find that the zinc finger portion of Disco binds in vitro to sequences located near the disco transcription unit, suggesting that Disco autoregulates its transcription in the optic lobe primordium by direct binding to a regulatory element in its own promoter.