Miracle-dz/UniFAR-FTUs huggingface dataset:
A Role for ZO-1 and PLEKHA7 in Recruiting Paracingulin to Tight and Adherens Junctions of Epithelial Cells*

Paracingulin is a 160-kDa protein localized in the cytoplasmic region of epithelial tight and adherens junctions, where it regulates RhoA and Rac1 activities by interacting with guanine nucleotide exchange factors. Here, we investigate the molecular mechanisms that control the recruitment of paracingulin to cell-cell junctions. We show that paracingulin forms a complex with the tight junction protein ZO-1, and the globular head domain of paracingulin interacts directly with ZO-1 through an N-terminal region containing a conserved ZIM (ZO-1-Interaction-Motif) sequence. Recruitment of paracingulin to cadherin-based cell-cell junctions in Rat1 fibroblasts requires the ZIM-containing region, whereas in epithelial cells removal of this region decreases the junctional localization of paracingulin at tight junctions but not at adherens junctions. Depletion of ZO-1, but not ZO-2, reduces paracingulin accumulation at tight junctions. A yeast two-hybrid screen identifies both ZO-1 and the adherens junction protein PLEKHA7 as paracingulin-binding proteins. Paracingulin forms a complex with PLEKHA7 and its interacting partner p120ctn, and the globular head domain of paracingulin interacts directly with a central region of PLEKHA7. Depletion of PLEKHA7 from Madin-Darby canine kidney cells results in the loss of junctional localization of paracingulin and a decrease in its expression. In summary, we characterize ZO-1 and PLEKHA7 as paracingulin-interacting proteins that are involved in its recruitment to epithelial tight and adherens junctions, respectively.
Identiﬁcation and characterization of a novel botulinum neurotoxin

Botulinum neurotoxins are known to have seven serotypes (BoNT/A–G). Here we report a new BoNT serotype, tentatively named BoNT/X, which has the lowest sequence identity with other BoNTs and is not recognized by antisera against known BoNTs. Similar to BoNT/B/D/F/G, BoNT/X cleaves vesicle-associated membrane proteins (VAMP) 1, 2 and 3, but at a novel site (Arg66-Ala67 in VAMP2). Remarkably, BoNT/X is the only toxin that also cleaves non-canonical substrates VAMP4, VAMP5 and Ykt6. To validate its activity, a small amount of full-length BoNT/X was assembled by linking two non-toxic fragments using a transpeptidase (sortase). Assembled BoNT/X cleaves VAMP2 and VAMP4 in cultured neurons and causes ﬂaccid paralysis in mice. Thus, BoNT/X is a novel BoNT with a unique substrate proﬁle. Its discovery posts a challenge to develop effective countermeasures, provides a novel tool for studying intracellular membrane trafﬁcking, and presents a new potential therapeutic toxin for modulating secretions in cells.
Antibacterial activity of dentine and pulp extracellular matrix extracts.

AIM To determine whether extracellular matrix (ECM) preparations from pulp (pECM) and dentine (dECM) possess antimicrobial activity. METHODOLOGY Dentine and pulp ECM preparations were isolated with 10% ethylenediaminetetraacetic acid (EDTA), pH 7.2 and sequential use of 0.5mol L(-1) NaCl, pH 11.7 and 0.1mol L(-1) tartaric acid, pH 2.0, respectively, with protease inhibitor inclusion throughout. Antimicrobial activity against Streptococcus mutans, Streptococcus oralis and Enterococcus faecalis was assessed using turbidity as a measure of bacteria growth. The cytotoxicity of the extracts on primary pulp cells was also determined by lactate dehydrogenase (LDH) release. Statistical analysis of data was performed using paired student's t-tests. RESULTS Extracellular matrix extracts from the pulp and dentine showed antibacterial activity against three types of anaerobic bacteria associated with dental disease (P< 0.05). The ECM extracts demonstrated no significant cytotoxic effect on pulpal cells at the concentrations used for antibacterial activity. CONCLUSIONS The bacteriostatic antibacterial activity of pECM and dECM indicates that the release of these matrix molecules from pulp and dentine may contribute to defence responses during dental disease, treatment and repair.
Contiguous gene syndrome due to a maternally inherited 8.41 Mb distal deletion of chromosome band Xp22.3 in a boy with short stature, ichthyosis, epilepsy, mental retardation, cerebral cortical heterotopias and Dandy–Walker malformation

Microdeletions of Xp22.3 are associated with contiguous gene syndromes, the extent and nature of which depend on the genes encompassed by the deletion. Common symptoms include ichthyosis, mental retardation and hypogonadism. We report on a boy with short stature, ichthyosis, severe mental retardation, cortical heterotopias and Dandy–Walker malformation. The latter two abnormalities have so far not been reported in terminal Xp deletions. MLPA showed deletion of SHOX and subsequent analysis using FISH and SNP‐arrays revealed that the patient had an 8.41 Mb distal deletion of chromosome region Xp22.31 → Xpter. This interval contains several genes whose deletion can partly explain our patient's phenotype. His cortical heterotopias and DWM suggest that a gene involved in brain development may be in the deleted interval, but we found no immediately obvious candidates. Interestingly, further analysis of the family revealed that the patient had inherited his deletion from his mother, who has a mos 46,X,del(X)(p22)/45,X/46, XX karyotype. © 2008 Wiley‐Liss, Inc.
Ten-year follow-up observations of periapical and endodontic status in a Danish population.

AIM To describe changes in periapical and endodontic status in a general adult Danish population examined in 1997, 2003 and 2008. METHODOLOGY The study provides longitudinal, observational information on endodontic status for 327 individuals who participated in three consecutive full-mouth radiographic examinations performed with an approximately 5-year interval. The collected data allow changes to be investigated within and between age-cohorts. RESULTS The median number of teeth declined slightly, but the number of individuals with apical periodontitis (AP) and with root filled teeth increased. The number of teeth with AP and of teeth with root fillings increased, whereas the number of root filled teeth with AP was almost stable. The proportion of individuals who had no AP and the proportion of individuals who had no root fillings decreased during the 10-year observation period. The proportion of teeth with AP/individual increased steadily with age, unaffected by the different time periods. Differences were found between the age groups for the proportion of root filled teeth/individual where the younger age-cohorts had root fillings performed later in life and at a slower rate. In the old age-cohort, the proportion of lost teeth/individual was much larger and showed a stronger increase with age, than in the younger age-cohorts. CONCLUSION The younger age-cohorts will probably never reach the proportion of root fillings or lost teeth seen in the oldest age-cohort. However, it is disturbing that the proportion of teeth with AP/individual increased steadily with age, completely unaffected by the different time periods.
Visual Insight into How Low pH Alone Can Induce Actin-severing Ability in Gelsolin under Calcium-free Conditions

Gelsolin is a key actin cytoskeleton-modulating protein primarily regulated by calcium and phosphoinositides. In addition, low pH has also been suggested to activate gelsolin in the absence of Ca2+ ions, although no structural insight on this pathway is available except for a reported decrement in its diffusion coefficient at low pH. We also observed ∼1.6-fold decrease in the molecular mobility of recombinant gelsolin when buffer pH was lowered from 9 to 5. Analysis of the small angle x-ray scattering data collected over the same pH range indicated that the radius of gyration and maximum linear dimension of gelsolin molecules increased from 30.3 to 34.1 Å and from 100 to 125 Å, respectively. Models generated for each dataset indicated that similar to the Ca2+-induced process, low pH also promotes unwinding of this six-domain protein but only partially. It appeared that pH is able to induce extension of the G1 domain from the rest of the five domains, whereas the Ca2+-sensitive latch between G2 and G6 domains remains closed. Interestingly, increasing the free Ca2+ level to merely ∼40 nm, the partially open pH 5 shape “sprung open” to a shape seen earlier for this protein at pH 8 and 1 mm free Ca2+. Also, pH alone could induce a shape where the g3-g4 linker of gelsolin was open when we truncated the C-tail latch from this protein. Our results provide insight into how under physiological conditions, a drop in pH can fully activate the F-actin-severing shape of gelsolin with micromolar levels of Ca2+ available.
Role of Novel Dimeric Photosystem II (PSII)-Psb27 Protein Complex in PSII Repair*

The multisubunit membrane protein complex Photosystem II (PSII) catalyzes one of the key reactions in photosynthesis: the light-driven oxidation of water. Here, we focus on the role of the Psb27 assembly factor, which is involved in biogenesis and repair after light-induced damage of the complex. We show that Psb27 is essential for the survival of cyanobacterial cells grown under stress conditions. The combination of cold stress (30 °C) and high light stress (1000 μmol of photons × m−2 × s−1) led to complete inhibition of growth in a Δpsb27 mutant strain of the thermophilic cyanobacterium Thermosynechococcus elongatus, whereas wild-type cells continued to grow. Moreover, Psb27-containing PSII complexes became the predominant PSII species in preparations from wild-type cells grown under cold stress. Two different PSII-Psb27 complexes were isolated and characterized in this study. The first complex represents the known monomeric PSII-Psb27 species, which is involved in the assembly of PSII. Additionally, a novel dimeric PSII-Psb27 complex could be allocated in the repair cycle, i.e. in processes after inactivation of PSII, by 15N pulse-label experiments followed by mass spectrometry analysis. Comparison with the corresponding PSII species from Δpsb27 mutant cells showed that Psb27 prevented the release of manganese from the previously inactivated complex. These results indicate a more complex role of the Psb27 protein within the life cycle of PSII, especially under stress conditions.
Recessive developmental delay, small stature, microcephaly and brain calcifications with locus on chromosome 2

Two interrelated Omani families are described with eight children manifesting a genetic disorder with widespread brain calcifications. Brain imaging showed extensive scattered calcifications of basal ganglia and cortex, suggesting possible Aicardi–Goutieres syndrome (AGS) or Coats' Plus syndrome. However, the clinical features in the present families diverge substantially from these two conditions. Growth delay, mild developmental delay, and poor school performance were present in all affected individuals, but progressive deterioration of neurological function was not apparent, nor were there significant cortical whitematter disease or retinopathy. Genome‐wide linkage and fine‐mapping analyses of the extended family members and affected individuals indicate a genetic locus for this disorder on Chromosome 2 with a LOD score of 6.17. The Chromosome 2 locus is novel and the clinical presentation displays features distinguishing the condition from either Coats' or AGS, making this a new variant or possibly a new disorder of inherited brain calcification. © 2009 Wiley‐Liss, Inc.
Runx2 Protein Expression Utilizes the Runx2 P1 Promoter to Establish Osteoprogenitor Cell Number for Normal Bone Formation*

The Runt-related transcription factor, Runx2, is essential for osteogenesis and is controlled by both distal (P1) and proximal (P2) promoters. To understand Runx2 function requires determination of the spatiotemporal activity of P1 and P2 to Runx2 protein production. We generated a mouse model in which the P1-derived transcript was replaced with a lacZ reporter allele, resulting in loss of P1-derived protein while simultaneously allowing discrimination between the activities of the two promoters. Loss of P1-driven expression causes developmental defects with cleidocranial dysplasia-like syndromes that persist in the postnatal skeleton. P1 activity is robust in preosteogenic mesenchyme and at the onset of bone formation but decreases as bone matures. Homozygous Runx2-P1lacZ/lacZ mice have a normal life span but exhibit severe osteopenia and compromised bone repair in adult mice because of osteoblastic defects and not increased osteoclastic resorption. Gene expression profiles of bone, immunohistochemical studies, and ex vivo differentiation using calvarial osteoblasts and marrow stromal cells identified mechanisms for the skeletal phenotype. The findings indicate that P1 promoter activity is necessary for generating a threshold level of Runx2 protein to commit sufficient osteoprogenitor numbers for normal bone formation. P1 promoter function is not compensated via the P2 promoter. However, the P2 transcript with compensatory mechanisms from bone morphogenetic protein (BMP) and Wnt signaling is adequate for mineralization of the bone tissue that does form. We conclude that selective utilization of the P1 and P2 promoters enables the precise spatiotemporal expression of Runx2 necessary for normal skeletogenesis and the maintenance of bone mass in the adult.
Coronavirus Infection Induces DNA Replication Stress Partly through Interaction of Its Nonstructural Protein 13 with the p125 Subunit of DNA Polymerase δ

Perturbation of cell cycle regulation is a characteristic feature of infection by many DNA and RNA viruses, including Coronavirus infectious bronchitis virus (IBV). IBV infection was shown to induce cell cycle arrest at both S and G2/M phases for the enhancement of viral replication and progeny production. However, the underlying mechanisms are not well explored. In this study we show that activation of cellular DNA damage response is one of the mechanisms exploited by Coronavirus to induce cell cycle arrest. An ATR-dependent cellular DNA damage response was shown to be activated by IBV infection. Suppression of the ATR kinase activity by chemical inhibitors and siRNA-mediated knockdown of ATR reduced the IBV-induced ATR signaling and inhibited the replication of IBV. Furthermore, yeast two-hybrid screens and subsequent biochemical and functional studies demonstrated that interaction between Coronavirus nsp13 and DNA polymerase δ induced DNA replication stress in IBV-infected cells. These findings indicate that the ATR signaling activated by IBV replication contributes to the IBV-induced S-phase arrest and is required for efficient IBV replication and progeny production.
Should early enteral nutrition be used in the trauma intensive care unit?

Critical illness, sepsis, surgery, multi-organ failure and haemodynamic instability are conditions that are associated with feeding inadequacies and nutritional challenges. Conversely, meeting nutritional requirements and providing optimal nutrition are associated with an improved outcome. Since these facts have been reported repeatedly, why then are we still faced with study results on suboptimal feeding?
Unexpected diagnosis of 45,X/47,XX,+18 mosaicism in a girl with mild phenotype

Unexpected Diagnosis of 45,X/47,XX,þ18 Mosaicism in a Girl With Mild Phenotype Caroline Schluth-Bolard, Damien Sanlaville, Audrey Labalme, Marianne Till, Laurence Michel-Calemard, Azim Rafat, Marie-Th er ese Zabot, Marc Nicolino, Laurent Guibaud, and Patrick Edery* Service de Cytog en etique Constitutionnelle, Hospices Civils de Lyon, Bron Cedex, France Facult e de M edecine Lyon Nord, Universit e Claude Bernard, Lyon, France Endocrinologie Mol eculaire et Maladies Rares, Hospices Civils de Lyon, Bron Cedex, France Centre de Biotechnologie Cellulaire, Hospices Civils de Lyon, Bron Cedex, France Service d’Endocrinologie P ediatrique, Hospices Civils de Lyon, Bron Cedex, France D epartement d’Imagerie P ediatrique et Fœtale, Hospices Civils de Lyon, Bron Cedex, France
Recent developments in biologic therapies for the treatment of patients with systemic lupus erythematosus

SLE has a complex pathogenesis, and multiple therapeutic targets have been discovered in recent years. In spite of belimumab being approved by the US Food and Drug Administration and the widespread use of rituximab, there have been many failed attempts to treat SLE successfully using biologic agents. In this review, we consider newer biologic approaches that might offer the hope of improving the outcome of SLE patients. These include the fully humanized anti-CD20 mAbs, PEGylated anti-CD40L, IFNα inhibitors, rigerimod and immune complexes blockade.
Evolutionary Remodeling of βγ-Crystallins for Domain Stability at Cost of Ca2+ Binding*

The topologically similar βγ-crystallins that are prevalent in all kingdoms of life have evolved for high innate domain stability to perform their specialized functions. The evolution of stability and its control in βγ-crystallins that possess either a canonical (mostly from microorganisms) or degenerate (principally found in vertebrate homologues) Ca2+-binding motif is not known. Using equilibrium unfolding of βγ-crystallin domains (26 wild-type domains and their mutants) in apo- and holo-forms, we demonstrate the presence of a stability gradient across these members, which is attained by the choice of residues in the (N/D)(N/D)XX(S/T)S Ca2+-binding motif. The occurrence of a polar, hydrophobic, or Ser residue at the 1st, 3rd, or 5th position of the motif is likely linked to a higher domain stability. Partial conversion of a microbe-type domain (with a canonical Ca2+-binding motif) to a vertebrate-type domain (with a degenerate Ca2+-binding motif) by mutating serine to arginine/lysine disables the Ca2+-binding but significantly augments its stability. Conversely, stability is compromised when arginine (in a vertebrate-type disabled domain) is replaced by serine (as a microbe type). Our results suggest that such conversions were acquired as a strategy for desired stability in vertebrate members at the cost of Ca2+-binding. In a physiological context, we demonstrate that a mutation such as an arginine to serine (R77S) mutation in this motif of γ-crystallin (partial conversion to microbe-type), implicated in cataracts, decreases the domain stability. Thus, this motif acts as a “central tuning knob” for innate as well as Ca2+-induced gain in stability, incorporating a stability gradient across βγ-crystallin members critical for their specialized functions.
Ubiquitin Ligase Substrate Identification through Quantitative Proteomics at Both the Protein and Peptide Levels

Background: Identification of ubiquitin ligase substrates remains an unmet challenge. Results: Two proteomic strategies were used to identify novel substrates of the E3 ligase HRD1. Conclusion: These methods identified populations of substrates enriched for potential targets of endoplasmic reticulum-associated degradation. Significance: This approach should be broadly useful for E3 ligase substrate identification, and the identified substrates provide insight into the role of HRD1 in disease. Protein ubiquitination is a key regulatory process essential to life at a cellular level; significant efforts have been made to identify ubiquitinated proteins through proteomics studies, but the level of success has not reached that of heavily studied post-translational modifications, such as phosphorylation. HRD1, an E3 ubiquitin ligase, has been implicated in rheumatoid arthritis, but no disease-relevant substrates have been identified. To identify these substrates, we have taken both peptide and protein level approaches to enrich for ubiquitinated proteins in the presence and absence of HRD1. At the protein level, a two-step strategy was taken using cells expressing His6-tagged ubiquitin, enriching proteins first based on their ubiquitination and second based on the His tag with protein identification by LC-MS/MS. Application of this method resulted in identification and quantification of more than 400 ubiquitinated proteins, a fraction of which were found to be sensitive to HRD1 and were therefore deemed candidate substrates. In a second approach, ubiquitinated peptides were enriched after tryptic digestion by peptide immunoprecipitation using an antibody specific for the diglycine-labeled internal lysine residue indicative of protein ubiquitination, with peptides and ubiquitination sites identified by LC-MS/MS. Peptide immunoprecipitation resulted in identification of over 1800 ubiquitinated peptides on over 900 proteins in each study, with several proteins emerging as sensitive to HRD1 levels. Notably, significant overlap exists between the HRD1 substrates identified by the protein-based and the peptide-based strategies, with clear cross-validation apparent both qualitatively and quantitatively, demonstrating the effectiveness of both strategies and furthering our understanding of HRD1 biology.
TRADICINIO INFORMUOTO ASMENS SUTIKIMO PRITAIKYMO MOKSLINIAMS BIOBANKAMS LIETUVOJE GALIMYBĖS

Siame straipsnyje siekiama įvertinti medicinos praktikoje isvystyto informuoto asmens sutikimo pritaikymo problematiką mokslinių biobankų veikloje, tai yra, paimant, tvarkant ir atliekant tyrimus su is žmogaus gauta biologine medžiaga (organais ar jų dalimis, audiniais, ląstelėmis ar jų komponentais), su sia medžiaga susijusia medicinine ar kita asmenį identifikuojancia informacija. Straipsnyje analizuojama, ar nežinant mokslinio tyrimo tikslo, jo trukmės ir galimų padarinių, bei galimo kito, nei buvo isreikstas sutikimas, panaudojimo tikslo asmuo gali būti laikomas tinkamai sutikes dalyvauti biobanko tyrimuose. Sis klausimas yra ypac problemiskas analizuojant aktyviąją teisės žinoti apie savo sveikatą puse, t. y. kai asmuo renkasi aktyvų domėjimąsi savo sveikata, nori gauti informacijos. Kartu sprendžiama, ar asmuo turi teise autonomiskai prisiimti riziką gaudamas ribotą informaciją is biobankų (tyrėjų), ir jeigu taip, tai kokiais teisiniais būdais būtų galima sių teisinių rizikų isvengti arba jas sumažinti. This article aims to assess the problematic of the applicablity of the informed consent, which was developed in the clinical practice, for the practice of the research biobanks, that is, taking, managing and conducting research with human biological material (organs or its parts, tissues, cells or their components), linked medical data or other informacion, which identifies the person. This article examines whether the lack of knowledge about the research objectives, duration of the research projects and possible consequences, as well as possible other, than expressed by the consent, purpose of use of such material, allows us to deem that the person has duly agreed to participate in biobank research. This question is specifically problematical, when analyzing the actine side of the rigzt to know about self health, i.e., when the person chooses the active interest in his health, wants to get the information. It is also being analyzed whether the person has the right to autonomously take the risk of receiving limited information from biobanks, and, if so, what legal safeguards would allow to avoid these legal risks or reduce them.