Huntington’s Disease protein huntingtin associates with its own mRNA

Background: The Huntington's disease (HD) protein huntingtin (Htt) plays a role in multiple cellular pathways. Deregulation of one or more of these pathways by the mutant Htt protein has been suggested to contribute to the disease pathogenesis. Our recent discovery-based proteomics studies have uncovered RNA binding proteins and translation factors associated with the endogenous Htt protein purified from mouse brains, suggesting a potential new role for Htt in RNA transport and translation. Objective: To investigate how Htt might affect RNA metabolism we set out to purify and analyze RNA associated with Htt. Methods: RNA was extracted from immunopurified Htt-containing protein complexes and analyzed by microarrays and RNA-Seq. Results: Surprisingly, the most enriched mRNA that co-purified with Htt was Htt mRNA itself. The association of Htt protein and Htt mRNA was detected independent of intact ribosomes suggesting that it is not an RNA undergoing translation. Furthermore, we identified the recently reported mis-spliced Htt mRNA encoding a truncated protein comprised of exon 1 and a portion of the downstream intron in the immunoprecipitates containing mutant Htt protein. We show that Htt protein co-localizes with Htt mRNA and that wild-type Htt reduces expression of a reporter construct harboring the Htt 3' UTR. Conclusions: HD protein is found in a complex with its own mRNA and RNA binding proteins and translation factors. Htt may be involved in modulating its expression through post-transcriptional pathways. It is possible that Htt shares mechanistic properties similar to RNA binding proteins such as TDP-43 and FUS implicated in other neurodegenerative diseases

LgR5 expression and cancer stem cell hypothesis: clue to define the true origin of esophageal adenocarcinomas with and without Barrett's Esophagus?

<p>Abstract</p> <p>Background</p> <p>Investigation of the expression of an intestinal stem cell marker in esophageal adenocarcinomas (EAC) with and without Barrett's Esophagus (BE), with respect to a cancer stem cell (CSC) hypothesis.</p> <p>Materials and methods</p> <p>Expression of a putative intestinal stem cell marker LgR5 was analyzed in esophageal cancer specimen (n = 70: 41 EAC with BE, 19 EAC without BE, and n = 10 esophageal squamous-cell carcinomas, ESCC) and in the adenocarcinoma cell line OE-33. Ki-67 and Cdx-2 were co-labelled with LgR5 in double staining experiments. Immunhistochemical expression results were confirmed by RT-PCR and correlated with tumor stage and five-year survival rates.</p> <p>Results</p> <p>LgR5was found expressed in 35 of 41 (85%) EAC with BE and in 16 of 19 (81%) EAC without BE. By contrast, LgR5 was not found to be expressed in ESCC. Quantification of immunolabeling showed 15% LgR5+ cells in EAC with BE, 32% LgR5+ cells in adjacent BE and 13% in EAC without BE. Immunofluorescence double staining experiments with LgR5 and Ki-67 revealed a subpopulation (~5%) of proliferating LgR+/Ki-67+ cells. On mRNA-level, expression of LgR5 was higher in BE in comparison to EAC (p = 0.0159). High levels of LgR5 expression in BE associated EAC were associated with poorer survival in univariate analysis.</p> <p>Conclusion</p> <p>The stem cell marker LgR5 is expressed in EAC, irrespective of association with BE, and appears to have negative impact on survival. The subset of proliferating LgR5+ cells (<5%) might resemble rapidly cycling CSCs, which needs to be substantiated in further investigations.</p

The Spatial Distribution of LGR5+ Cells Correlates With Gastric Cancer Progression

In this study we tested the prevalence, histoanatomical distribution and tumour biological significance of the Wnt target protein and cancer stem cell marker LGR5 in tumours of the human gastrointestinal tract. Differential expression of LGR5 was studied on transcriptional (real-time polymerase chain reaction) and translational level (immunohistochemistry) in malignant and corresponding non-malignant tissues of 127 patients comprising six different primary tumour sites, i.e. oesophagus, stomach, liver, pancreas, colon and rectum. The clinico-pathological significance of LGR5 expression was studied in 100 patients with gastric carcinoma (GC). Non-neoplastic tissue usually harboured only very few scattered LGR5+ cells. The corresponding carcinomas of the oesophagus, stomach, liver, pancreas, colon and rectum showed significantly more LGR5+ cells as well as significantly higher levels of LGR5-mRNA compared with the corresponding non-neoplastic tissue. Double staining experiments revealed a coexpression of LGR5 with the putative stem cell markers CD44, Musashi-1 and ADAM17. Next we tested the hypothesis that the sequential changes of gastric carcinogenesis, i.e. chronic atrophic gastritis, intestinal metaplasia and invasive carcinoma, are associated with a reallocation of the LGR5+ cells. Interestingly, the spatial distribution of LGR5 changed: in non-neoplastic stomach mucosa, LGR5+ cells were found predominantly in the mucous neck region; in intestinal metaplasia LGR5+ cells were localized at the crypt base, and in GC LGR5+ cells were present at the luminal surface, the tumour centre and the invasion front. The expression of LGR5 in the tumour centre and invasion front of GC correlated significantly with the local tumour growth (T-category) and the nodal spread (N-category). Furthermore, patients with LGR5+ GCs had a shorter median survival (28.0±8.6 months) than patients with LGR5− GCs (54.5±6.3 months). Our results show that LGR5 is differentially expressed in gastrointestinal cancers and that the spatial histoanatomical distribution of LGR5+ cells has to be considered when their tumour biological significance is sought

LGR6 Is a High Affinity Receptor of R-Spondins and Potentially Functions as a Tumor Suppressor

BACKGROUND: LGR6 (leucine-rich repeat containing, G protein-coupled receptor 6) is a member of the rhodopsin-like seven transmembrane domain receptor superfamily with the highest homology to LGR4 and LGR5. LGR6 was found as one of the novel genes mutated in colon cancer through total exon sequencing and its promoter region is hypermethylated in 20-50% of colon cancer cases. In the skin, LGR6 marks a population of stem cells that can give rise to all cell lineages. Recently, we and others demonstrated that LGR4 and LGR5 function as receptors of R-spondins to potentiate Wnt/β-catenin signaling. However, the binding affinity and functional response of LGR6 to R-spondins, and the activity of colon cancer mutants of LGR6 have not been determined. PRINCIPAL FINDINGS: We found that LGR6 also binds and responds to R-spondins 1-3 with high affinity to enhance Wnt/β-catenin signaling through increased LRP6 phosphorylation. Similar to LGR4 and LGR5, LGR6 is not coupled to heterotrimeric G proteins or to β-arrestin following R-spondin stimulation. Functional and expression analysis of three somatic mutations identified in colon cancer samples indicates that one mutant fails to bind and respond to R-spondin (loss-of-function), but the other two have no significant effect on receptor function. Overexpression of wild-type LGR6 in HeLa cells leads to increased cell migration following co-treatment with R-spondin1 and Wnt3a when compared to vector control cells or cells overexpressing the loss-of-function mutant. CONCLUSIONS: LGR6 is a high affinity receptor for R-spondins 1-3 and potentially functions as a tumor suppressor despite its positive effect on Wnt/β-catenin signaling

Gene Expression Profiling of Embryonic Human Neural Stem Cells and Dopaminergic Neurons from Adult Human Substantia Nigra

Neural stem cells (NSC) with self-renewal and multipotent properties serve as an ideal cell source for transplantation to treat neurodegenerative insults such as Parkinson's disease. We used Agilent's and Illumina Whole Human Genome Oligonucleotide Microarray to compare the genomic profiles of human embryonic NSC at a single time point in culture, and a multicellular tissue from postmortem adult substantia nigra (SN) which are rich in dopaminergic (DA) neurons. We identified 13525 up-regulated genes in both cell types of which 3737 (27.6%) genes were up-regulated in the hENSC, 4116 (30.4%) genes were up-regulated in the human substantia nigra dopaminergic cells, and 5672 (41.93%) were significantly up-regulated in both cell population. Careful analysis of the data that emerged using DAVID has permitted us to distinguish several genes and pathways that are involved in dopaminergic (DA) differentiation, and to identify the crucial signaling pathways that direct the process of differentiation. The set of genes expressed more highly at hENSC is enriched in molecules known or predicted to be involved in the M phase of the mitotic cell cycle. On the other hand, the genes enriched in SN cells include a different set of functional categories, namely synaptic transmission, central nervous system development, structural constituents of the myelin sheath, the internode region of axons, myelination, cell projection, cell somata, ion transport, and the voltage-gated ion channel complex. Our results were also compared with data from various databases, and between different types of arrays, Agilent versus Illumina. This approach has allowed us to confirm the consistency of our obtained results for a large number of genes that delineate the phenotypical differences of embryonic NSCs, and SN cells

The HIV-1 Integrase α4-Helix Involved in LTR-DNA Recognition Is also a Highly Antigenic Peptide Element

Monoclonal antibodies (MAbas) constitute remarkable tools to analyze the relationship between the structure and the function of a protein. By immunizing a mouse with a 29mer peptide (K159) formed by residues 147 to 175 of the HIV-1 integrase (IN), we obtained a monoclonal antibody (MAba4) recognizing an epitope lying in the N-terminal portion of K159 (residues 147–166 of IN). The boundaries of the epitope were determined in ELISA assays using peptide truncation and amino acid substitutions. The epitope in K159 or as a free peptide (pep-a4) was mostly a random coil in solution, while in the CCD (catalytic core domain) crystal, the homologous segment displayed an amphipathic helix structure (α4-helix) at the protein surface. Despite this conformational difference, a strong antigenic crossreactivity was observed between pep-a4 and the protein segment, as well as K156, a stabilized analogue of pep-a4 constrained into helix by seven helicogenic mutations, most of them involving hydrophobic residues. We concluded that the epitope is freely accessible to the antibody inside the protein and that its recognition by the antibody is not influenced by the conformation of its backbone and the chemistry of amino acids submitted to helicogenic mutations. In contrast, the AA →Glu mutations of the hydrophilic residues Gln148, Lys156 and Lys159, known for their interactions with LTRs (long terminal repeats) and inhibitors (

Real-space collapse of a polariton condensate

Microcavity polaritons are two-dimensional bosonic fluids with strong nonlinearities, composed of coupled photonic and electronic excitations. In their condensed form, they display quantum hydrodynamic features similar to atomic Bose–Einstein condensates, such as long-range coherence, superfluidity and quantized vorticity. Here we report the unique phenomenology that is observed when a pulse of light impacts the polariton vacuum: the fluid which is suddenly created does not splash but instead coheres into a very bright spot. The real-space collapse into a sharp peak is at odd with the repulsive interactions of polaritons and their positive mass, suggesting that an unconventional mechanism is at play. Our modelling devises a possible explanation in the self-trapping due to a local heating of the crystal lattice, that can be described as a collective polaron formed by a polariton condensate. These observations hint at the polariton fluid dynamics in conditions of extreme intensities and ultrafast times

Ultra-low-power hybrid light-matter solitons.

New functionalities in nonlinear optics will require systems with giant optical nonlinearity as well as compatibility with photonic circuit fabrication techniques. Here we introduce a platform based on strong light-matter coupling between waveguide photons and quantum-well excitons. On a sub-millimetre length scale we generate picosecond bright temporal solitons at a pulse energy of only 0.5 pJ. From this we deduce a nonlinear refractive index three orders of magnitude larger than in any other ultrafast system. We study both temporal and spatio-temporal nonlinear effects and observe dark-bright spatio-temporal polariton solitons. Theoretical modelling of soliton formation in the strongly coupled system confirms the experimental observations. These results show the promise of our system as a high speed, low power, integrated platform for physics and devices based on strong interactions between photons