Stochastic state transitions give rise to phenotypic equilibrium in populations of cancer cells

Cancer cells within individual tumors often exist in distinct phenotypic states that differ in functional attributes. While cancer cell populations typically display distinctive equilibria in the proportion of cells in various states, the mechanisms by which this occurs are poorly understood. Here, we study the dynamics of phenotypic proportions in human breast cancer cell lines. We show that subpopulations of cells purified for a given phenotypic state return towards equilibrium proportions over time. These observations can be explained by a Markov model in which cells transition stochastically between states. A prediction of this model is that, given certain conditions, any subpopulation of cells will return to equilibrium phenotypic proportions over time. A second prediction is that breast cancer stem-like cells arise de novo from non-stem-like cells. These findings contribute to our understanding of cancer heterogeneity and reveal how stochasticity in single-cell behaviors promotes phenotypic equilibrium in populations of cancer cells.     

Molecular architecture and connectivity of the budding yeast Mtw1 kinetochore complex

Kinetochores are large multiprotein complexes that connect centromeres to spindle microtubules in all eukaryotes. Among the biochemically distinct kinetochore complexes, the conserved four-protein Mtw1 complex is a central part of the kinetochore in all organisms. Here we present the biochemical reconstitution and characterization of the budding yeast Mtw1 complex. Direct visualization by electron microscopy revealed an elongated bilobed structure with a 25-nm-long axis. The complex can be assembled from two stable heterodimers consisting of Mtw1p-Nnf1p and Dsn1p-Nsl1p, and it interacts directly with the microtubule-binding Ndc80 kinetochore complex via the centromere-proximal Spc24/Spc25 head domain. In addition, we have reconstituted a partial Ctf19 complex and show that it directly associates with the Mtw1 complex in vitro. Ndc80 and Ctf19 complexes do not compete for binding to the Mtw1 complex, suggesting that Mtw1 can bridge the microtubule-binding components of the kinetochore to the inner centromere.     

Cohesin mediates chromatin interactions that regulate mammalian β-globin expression

The β-globin locus undergoes dynamic chromatin interaction changes in differentiating erythroid cells that are thought to be important for proper globin gene expression. However, the underlying mechanisms are unclear. The CCCTC-binding factor, CTCF, binds to the insulator elements at the 5' and 3' boundaries of the locus, but these sites were shown to be dispensable for globin gene activation. We found that, upon induction of differentiation, cohesin and the cohesin loading factor Nipped-B-like (Nipbl) bind to the locus control region (LCR) at the CTCF insulator and distal enhancer regions as well as at the specific target globin gene that undergoes activation upon differentiation. Nipbl-dependent cohesin binding is critical for long-range chromatin interactions, both between the CTCF insulator elements and between the LCR distal enhancer and the target gene. We show that the latter interaction is important for globin gene expression in vivo and in vitro. Furthermore, the results indicate that such cohesin-mediated chromatin interactions associated with gene regulation are sensitive to the partial reduction of Nipbl caused by heterozygous mutation. This provides the first direct evidence that Nipbl haploinsufficiency affects cohesin-mediated chromatin interactions and gene expression. Our results reveal that dynamic Nipbl/cohesin binding is critical for developmental chromatin organization and the gene activation function of the LCR in mammalian cells.     

Contribution of the Ly49E Natural Killer Receptor in the Immune Response to Plasmodium berghei Infection and Control of Hepatic Parasite Development

Natural killer (NK) cells have different roles in the host response against Plasmodium-induced malaria depending on the stage of infection. Liver NK cells have a protective role during the initial hepatic stage of infection by production of the T_H1-type cytokines IFN-γ and TNF-α. In the subsequent erythrocytic stage of infection, NK cells also induce protection through Th1-type cytokines but, in addition, may also promote development of cerebral malaria via CXCR3-induction on CD8⁺ T cells resulting in migration of these cells to the brain. We have recently shown that the regulatory Ly49E NK receptor is expressed on liver NK cells in particular. The main objective of this study was therefore to examine the role of Ly49E expression in the immune response upon Plasmodium berghei ANKA infection, for which we compared wild type (WT) to Ly49E knockout (KO) mice. We show that the parasitemia was higher at the early stage, i.e. at days 6–7 of Plasmodium berghei ANKA infection in Ly49E KO mice, which correlated with lower induction of CD69, IFN-γ and TNF-α in DX5⁻ liver NK cells at day 5 post-infection. At later stages, these differences faded. There was also no difference in the kinetics and the percentage of cerebral malaria development and in lymphocyte CXCR3 expression in WT versus Ly49E KO mice. Collectively, we show that the immune response against Plasmodium berghei ANKA infection is not drastically affected in Ly49E KO mice. Although NK cells play a crucial role in Plasmodium infection and Ly49E is highly expressed on liver NK cells, the Ly49E NK receptor only has a temporarily role in the immune control of this parasite.     

Current Practice of Public Involvement Activities in Biomedical Research and Innovation: A Systematic Qualitative Review

Background

A recent report from the British Nuffield Council on Bioethics associated ‘emerging biotechnologies’ with a threefold challenge: 1) uncertainty about outcomes, 2) diverse public views on the values and implications attached to biotechnologies and 3) the possibility of creating radical changes regarding societal relations and practices. To address these challenges, leading international institutions stress the need for public involvement activities (PIAs). The objective of this study was to assess the state of PIA reports in the field of biomedical research.

Methods

PIA reports were identified via a systematic literature search. Thematic text analysis was employed for data extraction.

Results

After filtering, 35 public consultation and 11 public participation studies were included in this review. Analysis and synthesis of all 46 PIA studies resulted in 6 distinguishable PIA objectives and 37 corresponding PIA methods. Reports of outcome translation and PIA evaluation were found in 9 and 10 studies respectively (20% and 22%). The paper presents qualitative details.

Discussion

The state of PIAs on biomedical research and innovation is characterized by a broad range of methods and awkward variation in the wording of objectives. Better comparability of PIAs might improve the translation of PIA findings into further policy development. PIA-specific reporting guidelines would help in this regard. The modest level of translation efforts is another pointer to the “deliberation to policy gap”. The results of this review could inform the design of new PIAs and future efforts to improve PIA comparability and outcome translation.     

Holy Smoke in Medieval Funerary Rites: Chemical Fingerprints of Frankincense in Southern Belgian Incense Burners

Frankincense, the oleogum resin from Boswellia sp., has been an early luxury good in both Western and Eastern societies and is particularly used in Christian funerary and liturgical rites. The scant grave goods in late medieval burials comprise laterally perforated pottery vessels which are usually filled with charcoal. They occur in most regions of western Europe and are interpreted as incense burners but have never been investigated with advanced analytical techniques. We herein present chemical and anthracological results on perforated funerary pots from 4 Wallonian sites dating to the 12–14^th century AD. Chromatographic and mass spectrometric analysis of lipid extracts of the ancient residues and comparison with extracts from four Boswellia species clearly evidence the presence of degraded frankincense in the former, based on characteristic triterpenoids, viz. boswellic and tirucallic acids, and their myriad dehydrated and oxygenated derivatives. Cembrane-type diterpenoids indicate B. sacra (southern Arabia) and B. serrata (India) as possible botanical origins. Furthermore, traces of juniper and possibly pine tar demonstrate that small amounts of locally available fragrances were mixed with frankincense, most likely to reduce its cost. Additionally, markers of ruminant fats in one sample from a domestic context indicate that this vessel was used for food preparation. Anthracological analysis demonstrates that the charcoal was used as fuel only and that no fragrant wood species were burned. The chars derived from local woody plants and were most likely recovered from domestic fires. Furthermore, vessel recycling is indicated by both contextual and biomarker evidence. The results shed a new light on funerary practices in the Middle Ages and at the same time reveal useful insights into the chemistry of burned frankincense. The discovery of novel biomarkers, namely Δ²-boswellic acids and a series of polyunsaturated and aromatic hydrocarbons, demonstrates the high potential for organic chemical analyses of incense residues.