Methylome profiling reveals functions and genes which are differentially methylated in serrated compared to conventional colorectal carcinoma.

Background: Serrated adenocarcinoma (SAC) is a recently recognized colorectal cancer (CRC) subtype accounting for 7.5–8.7 % of CRCs. It has been shown that SAC has a worse prognosis and different histological and molecular features compared to conventional carcinoma (CC) but, to date, there is no study analysing its methylome profile. Results: The methylation status of 450,000 CpG sites using the Infinium Human Methylation 450 BeadChip array was investigated in 103 colorectal specimens, including 39 SACs and 34 matched CCs, from Spanish and Finnish patients. Microarray data showed a higher representation of morphogenesis-, neurogenesis-, cytoskeleton- and vesicle transport-related functions and also significant differential methylation of 15 genes, including the iodothyronine deiodinase DIO3 and the forkhead family transcription factor FOXD2 genes which were validated at the CpG, mRNA and protein level using pyrosequencing, methylation-specific PCR, quantitative polymerase chain reaction (qPCR) and immunohistochemistry. A quantification study of the methylation status of CpG sequences in FOXD2 demonstrated a novel region controlling gene expression. Moreover, differences in these markers were also evident when comparing SAC with CRC showing molecular and histological features of high-level microsatellite instability. Conclusions: This methylome study demonstrates distinct epigenetic regulation patterns in SAC which are consistent to previous expression profile studies and that DIO3 and FOXD2 might be molecular targets for a specific histologyoriented treatment of CRC.post-print2306 K

CD94-NKG2A recognition of human leukocyte antigen (HLA)-E bound to an HLA class I leader sequence

The recognition of human leukocyte antigen (HLA)-E by the heterodimeric CD94-NKG2 natural killer (NK) receptor family is a central innate mechanism by which NK cells monitor the expression of other HLA molecules, yet the structural basis of this highly specific interaction is unclear. Here, we describe the crystal structure of CD94-NKG2A in complex with HLA-E bound to a peptide derived from the leader sequence of HLA-G. The CD94 subunit dominated the interaction with HLA-E, whereas the NKG2A subunit was more peripheral to the interface. Moreover, the invariant CD94 subunit dominated the peptide-mediated contacts, albeit with poor surface and chemical complementarity. This unusual binding mode was consistent with mutagenesis data at the CD94-NKG2A–HLA-E interface. There were few conformational changes in either CD94-NKG2A or HLA-E upon ligation, and such a “lock and key” interaction is typical of innate receptor–ligand interactions. Nevertheless, the structure also provided insight into how this interaction can be modulated by subtle changes in the peptide ligand or by the pairing of CD94 with other members of the NKG2 family. Differences in the docking strategies used by the NKG2D and CD94-NKG2A receptors provided a basis for understanding the promiscuous nature of ligand recognition by NKG2D compared with the fidelity of the CD94-NKG2 receptors

Admissions for hypoglycaemia after 35 weeks of gestation:perinatal predictors of cost of stay

Multivariate analysis of the clinicopathological and molecular factors associated with FOXD2 methylation (A); DIO3 and FOXD2 mRNA expression in hMSI-H tumoural and normal specimens (B) and external validation using TCGA database showing that MSI-H expressed less FOXD2 than MSI-L/MSS colon carcinomas (C)

Giving an Account of One’s Pain in the Anthropological Interview

In this paper, I analyze the illness stories narrated by a mother and her 13-year-old son as part of an ethnographic study of child chronic pain sufferers and their families. In examining some of the moral, relational and communicative challenges of giving an account of one’s pain, I focus on what is left out of some accounts of illness and suffering and explore some possible reasons for these elisions. Drawing on recent work by Judith Butler (Giving an Account of Oneself, 2005), I investigate how the pragmatic context of interviews can introduce a form of symbolic violence to narrative accounts. Specifically, I use the term “genre of complaint” to highlight how anthropological research interviews in biomedical settings invoke certain typified forms of suffering that call for the rectification of perceived injustices. Interview narratives articulated in the genre of complaint privilege specific types of pain and suffering and cast others into the background. Giving an account of one’s pain is thus a strategic and selective process, creating interruptions and silences as much as moments of clarity. Therefore, I argue that medical anthropologists ought to attend more closely to the institutional structures and relations that shape the production of illness narratives in interview encounters

Natural micropolymorphism in human leukocyte antigens provides a basis for genetic control of antigen recognition

Human leukocyte antigen (HLA) gene polymorphism plays a critical role in protective immunity, disease susceptibility, autoimmunity, and drug hypersensitivity, yet the basis of how HLA polymorphism influences T cell receptor (TCR) recognition is unclear. We examined how a natural micropolymorphism in HLA-B44, an important and large HLA allelic family, affected antigen recognition. T cell–mediated immunity to an Epstein-Barr virus determinant (EENLLDFVRF) is enhanced when HLA-B*4405 was the presenting allotype compared with HLA-B*4402 or HLA-B*4403, each of which differ by just one amino acid. The micropolymorphism in these HLA-B44 allotypes altered the mode of binding and dynamics of the bound viral epitope. The structure of the TCR–HLA-B*4405EENLLDFVRF complex revealed that peptide flexibility was a critical parameter in enabling preferential engagement with HLA-B*4405 in comparison to HLA-B*4402/03. Accordingly, major histocompatibility complex (MHC) polymorphism can alter the dynamics of the peptide-MHC landscape, resulting in fine-tuning of T cell responses between closely related allotypes

A green fluorescent protein containing a QFG tri-peptide chromophore: optical properties and X-Ray crystal structure

Rtms5 is an deep blue weakly fluorescent GFP-like protein (lambda(max)(Abs), 592 nm; lambda(max)(Em), 630nm; Phi(F), 0.004) that contains a (66)Gln-Tyr-Gly chromophore tripeptide sequence. We investigated the optical properties and structure of two variants, Rtms5(Y67F) and Rtms5(Y67F/H146S) in which the tyrosine at position 67 was substituted by a phenylalanine. Compared to the parent proteins the optical spectra for these new variants were significantly blue-shifted. Rtms5(Y67F) spectra were characterised by two absorbing species (lambda(max)(Abs), 440 nm and 513 nm) and green fluorescence emission (lambda(max)(Ex), 440 nm; lambda(max)(Em), 508 nm; Phi(F), 0.11), whilst Rtms5(Y67F/H146S) spectra were characterised by a single absorbing species (lambda(max)(Abs), 440 nm) and a relatively high fluorescence quantum yield (Phi(F), 0.75; lambda(max)(Ex), 440 nm; lambda(max)(Em), 508 nm). The fluorescence emissions of each variant were remarkably stable over a wide range of pH (3-11). These are the first GFP-like proteins with green emissions (500-520 nm) that do not have a tyrosine at position 67. The X-ray crystal structure of each protein was determined to 2.2 angstrom resolution and showed that the benzylidine ring of the chromophore, similar to the 4-hydroxybenzylidine ring of the Rtms5 parent, is non-coplanar and in the trans conformation. The results of chemical quantum calculations together with the structural data suggested that the 513 nm absorbing species in Rtms5(Y67F) results from an unusual form of the chromophore protonated at the acylimine oxygen. These are the first X-ray crystal structures for fluorescent proteins with a functional chromophore containing a phenylalanine at position 67

The chromophore model used for quantum chemical calculations.

<p>The chromophore model is truncated at a level consistent with earlier studies of acylimine-substituted FP chromophore models. The neutral unprotonated form is shown. The protonation sites for each of the three singly protonated forms are indicated.</p