Determination of the Subclonal Tumor Structure in Childhood Acute Myeloid Leukemia and Acral Melanoma by Next-Generation Sequencing

Molecular Biology - Intratumoral heterogeneity and clonal variability are among the central problems of clinical oncology, leading to resistance to therapy, relapse, and metastasis. High-throughput...     

The mechanobiology theory of the development of medical device-related pressure ulcers revealed through a cell-scale computational modeling framework

Biomechanics and Modeling in Mechanobiology - Pressure ulcers are localized sites of tissue damage which form due to the continuous exposure of skin and underlying soft tissues to sustained...     

Magnetic Fractionation and Proteomic Dissection of Cellular Organelles Occupied by the Late Replication Complexes of Semliki Forest Virus

Alphavirus replicase complexes are initially formed at the plasma membrane and are subsequently internalized by endocytosis. During the late stages of infection, viral replication organelles are represented by large cytopathic vacuoles, where replicase complexes bind to membranes of endolysosomal origin. In addition to viral components, these organelles harbor an unknown number of host proteins. In this study, a fraction of modified lysosomes carrying functionally intact replicase complexes was obtained by feeding Semliki Forest virus (SFV)-infected HeLa cells with dextran-covered magnetic nanoparticles and later magnetically isolating the nanoparticle-containing lysosomes. Stable isotope labeling with amino acids in cell culture combined with quantitative proteomics was used to reveal 78 distinct cellular proteins that were at least 2.5-fold more abundant in replicase complex-carrying vesicles than in vesicles obtained from noninfected cells. These host components included the RNA-binding proteins PCBP1, hnRNP M, hnRNP C, and hnRNP K, which were shown to colocalize with the viral replicase. Silencing of hnRNP M and hnRNP C expression enhanced the replication of SFV, Chikungunya virus (CHIKV), and Sindbis virus (SINV). PCBP1 silencing decreased SFV-mediated protein synthesis, whereas hnRNP K silencing increased this synthesis. Notably, the effect of hnRNP K silencing on CHIKV- and SINV-mediated protein synthesis was opposite to that observed for SFV. This study provides a new approach for analyzing the proteome of the virus replication organelle of positive-strand RNA viruses and helps to elucidate how host RNA-binding proteins exert important but diverse functions during positive-strand RNA viral infection.     

Molecular biogeography of red deer Cervus elaphus from eastern Europe: insights from mitochondrial DNA sequences

European red deer are known to show a conspicuous phylogeographic pattern with three distinct mtDNA lineages (western, eastern and North-African/Sardinian). The western lineage, believed to be indicative of a southwestern glacial refuge in Iberia and southern France, nowadays covers large areas of the continent including the British Isles, Scandinavia and parts of central Europe, while the eastern lineage is primarily found in southeast-central Europe, the Carpathians and the Balkans. However, large parts of central Europe and the whole northeast of the continent were not covered by previous analyses. To close this gap, we produced mtDNA control region sequences from more than 500 red deer from Denmark, Germany, Poland, Lithuania, Belarus, Ukraine and western Russia and combined our data with sequences available from earlier studies to an overall sample size of almost 1,100. Our results show that the western lineage extends far into the European east and is prominent in all eastern countries except for the Polish Carpathians, Ukraine and Russia where only eastern haplotypes occurred. While the latter may actually reflect the natural northward expansion of the eastern lineage after the last ice age, the present distribution of the western lineage in eastern Europe may in large parts be artificial and a result of translocations and reintroduction of red deer into areas where the species became extinct in historical times.     

Body size and coexistence in gamasid mites parasitic on small mammals: null model analyses at three hierarchical scales

We studied body size ratio in gamasid mites (Acari: Mesostigmata) parasitic on Palearctic small mammals at 3 hierarchical scales, namely infracommunities (an assemblage of mites harboured by an individual host), component communities (an assemblage of mites harboured by a host population), and compound communities (an assemblage of mites harboured by a host community). We used null models and asked a) whether body size distributions in these communities demonstrate non‐random patterns; b) whether these patterns indicate segregation or aggregation of body sizes of coexisting species; and c) whether patterns of body size distribution are scale‐dependent, that is, differ among infracommunities, component communities, and compound communities. In most mite assemblages, the observed pattern of body size distribution did not differ from that expected by chance. However, meta‐analyses demonstrated that component and compound communities of gamasid mites consistently demonstrated a tendency to reduced body size overlap, while we did not find any clear trend in mite body size distribution across infracommunities. We discuss reasons for scale‐dependence of body size distribution pattern in parasite communities and propose ecological and evolutionary mechanisms that allowed the reduced body size overlap in component and compound communities of ectoparasites to arise.     

Vertical nontransovarial transmission of Bartonella in fleas

Pathogens use diverse pathways to infect host populations by vertical and/or horizontal routes. Horizontal transmission of bacteria belonging to the Bartonella genus via haematophagous vectors is well known. Vertical transmission of Bartonella species was also suggested to occur but its routes remain to be unveiled. In a previous study, we showed the absence of transovarial transmission of Bartonella species OE 1‐1 in Xenopsylla ramesis fleas, and that fleas feeding on Bartonella‐positive jirds produced Bartonella‐positive gut voids. This current study aimed to investigate whether vertical nontransovarial transmission of Bartonella occurs in fleas. For this aim, the X. ramesis–Bartonella sp. OE 1‐1 model was used. Four groups of fleas including Bartonella‐positive and Bartonella‐negative female fleas and larval offspring had access to either Bartonella‐negative or Bartonella‐positive gut voids and faeces. Sixteen per cent of flea offspring that had access to Bartonella‐positive faeces and gut voids became Bartonella positive. Our findings demonstrate that Bartonella‐positive flea faeces and gut voids are proper infection sources for flea larvae and indicate that vertical nontransovarial transmission of bartonellae occurs in fleas. This information broadens our understanding of Bartonella transmission routes in flea vectors and enlightens pathways of bartonellae transmission and maintenance in flea populations in nature.     

Novel reference gene RPN1 for normalization of quantitative data in lung and kidney cancer

Quantitative methods of gene expression analysis in tumors require accurate data normalization, which allows comparison of different mRNA/cDNA samples with unknown concentration. For this purpose reference genes with stable expression level (such as GAPDH, ACTB, HPRT1, TBP) are used. The choice of appropriate reference genes is still actual because well-known reference genes are not suitable for certain cancer types frequently and their unreasonable use without additional tests lead to wrong conclusions. We have developed the bioinformatic approach and selected a new potential reference gene RPN1 for lung and kidney tumors. This gene is located at the long arm of chromosome 3. Our method includes mining of the dbEST and Oncomine databases and functional analysis of genes. The RPN1 was selected from 1500 candidate housekeeping genes. Using comparative genomic hybridization with NotI-microarrays we found no methylation, deletions and/or amplifications at the RPN1-containing locus in 56 non-small cell lung and 42 clear cell renal cancer samples. Using RT-qPCR we showed low variability of RPN1 mRNA level comparable to those of reference genes GAPDH and GUSB in lung and kidney cancer. The mRNA levels of two target genes coding hyalouronidases--HYAL1 and HYAL2--were estimated and normalized relative to pair RPN1--GAPDH genes for lung cancer and RPN1--GUSB for kidney cancer. These combinations were shown to be optimal for obtaining accurate and reproducible data. All obtained results allow us to suggest RPN1 as novel reference gene for quantitative data normalization in gene expression studies for lung and kidney cancers.