Mnt–Max to Myc–Max complex switching regulates cell cycle entry

The c-Myc oncoprotein is strongly induced during the G0 to S-phase transition and is an important regulator of cell cycle entry. In contrast to c-Myc, the putative Myc antagonist Mnt is maintained at a constant level during cell cycle entry. Mnt and Myc require interaction with Max for specific DNA binding at E-box sites, but have opposing transcriptional activities. Here, we show that c-Myc induction during cell cycle entry leads to a transient decrease in Mnt–Max complexes and a transient switch in the ratio of Mnt–Max to c-Myc–Max on shared target genes. Mnt overexpression suppressed cell cycle entry and cell proliferation, suggesting that the ratio of Mnt–Max to c-Myc–Max is critical for cell cycle entry. Furthermore, simultaneous Cre-Lox mediated deletion of Mnt and c-Myc in mouse embryo fibroblasts rescued the cell cycle entry and proliferative block caused by c-Myc ablation alone. These results demonstrate that Mnt-Myc antagonism plays a fundamental role in regulating cell cycle entry and proliferation

Gene expression of the insulin-like growth factors and their receptors in human neuroblastoma cell lines

Insulin-like growth factors (IGF) I and II are polypeptides with both growth-promoting and insulin-like metabolic effects. The developmentally specific expression of IGF I and II in the nervous system implies a role for these growth factors in neuronal growth and differentiation. In the present study, we analyzed IGF and IGF receptor mRNA transcripts from two related human neuroblastoma cell lines, SH-SY5Y and SK-N-SH. These cell lines provide a good in vitro model of neuronal development. Northern analysis of total RNA from each cell line revealed three IGF II mRNA transcripts (6.0, 4.8, and 1.8 kb), and one mRNA transcript each for the type I (11.0 kb) and type II (9.4 kb) IGF receptors. The size distribution of these multiple transcripts is similar to that found during normal human fetal development. These results establish both cell lines as good in vitro models for investigating the mechanisms which underly IGF gene expression during nervous system development.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/29806/1/0000152.pd

Sequential and Coordinated Actions of c-Myc and N-Myc Control Appendicular Skeletal Development

BACKGROUND: During limb development, chondrocytes and osteoblasts emerge from condensations of limb bud mesenchyme. These cells then proliferate and differentiate in separate but adjacent compartments and function cooperatively to promote bone growth through the process of endochondral ossification. While many aspects of limb skeletal formation are understood, little is known about the mechanisms that link the development of undifferentiated limb bud mesenchyme with formation of the precartilaginous condensation and subsequent proliferative expansion of chondrocyte and osteoblast lineages. The aim of this study was to gain insight into these processes by examining the roles of c-Myc and N-Myc in morphogenesis of the limb skeleton. METHODOLOGY/PRINCIPAL FINDINGS: To investigate c-Myc function in skeletal development, we characterized mice in which floxed c-Myc alleles were deleted in undifferentiated limb bud mesenchyme with Prx1-Cre, in chondro-osteoprogenitors with Sox9-Cre and in osteoblasts with Osx1-Cre. We show that c-Myc promotes the proliferative expansion of both chondrocytes and osteoblasts and as a consequence controls the process of endochondral growth and ossification and determines bone size. The control of proliferation by c-Myc was related to its effects on global gene transcription, as phosphorylation of the C-Terminal Domain (pCTD) of RNA Polymerase II, a marker of general transcription initiation, was tightly coupled to cell proliferation of growth plate chondrocytes where c-Myc is expressed and severely downregulated in the absence of c-Myc. Finally, we show that combined deletion of N-Myc and c-Myc in early limb bud mesenchyme gives rise to a severely hypoplastic limb skeleton that exhibits features characteristic of individual c-Myc and N-Myc mutants. CONCLUSIONS/SIGNIFICANCE: Our results show that N-Myc and c-Myc act sequentially during limb development to coordinate the expansion of key progenitor populations responsible for forming the limb skeleton

Active gas features in three HSC-SSP CAMIRA clusters revealed by high angular resolution analysis of MUSTANG-2 SZE and XXL X-ray observations

International audienceWe present results from simultaneous modelling of high angular resolution GBT/MUSTANG-2 90 GHz Sunyaev–Zel’dovich effect (SZE) measurements and XMM-XXL X-ray images of three rich galaxy clusters selected from the HSC-SSP Survey. The combination of high angular resolution SZE and X-ray imaging enables a spatially resolved multicomponent analysis, which is crucial to understand complex distributions of cluster gas properties. The targeted clusters have similar optical richnesses and redshifts, but exhibit different dynamical states in their member galaxy distributions: a single-peaked cluster, a double-peaked cluster, and a cluster belonging to a supercluster. A large-scale residual pattern in both regular Compton-parameter y and X-ray surface brightness distributions is found in the single-peaked cluster, indicating a sloshing mode. The double-peaked cluster shows an X-ray remnant cool core between two SZE peaks associated with galaxy concentrations. The temperatures of the two peaks reach ∼20–30 keV in contrast to the cool core component of ∼2 keV, indicating a violent merger. The main SZE signal for the supercluster is elongated along a direction perpendicular to the major axis of the X-ray core, suggesting a minor merger before core passage. The and y distributions are thus perturbed at some level, regardless of the optical properties. We find that the integrated Compton y parameter and the temperature for the major merger are boosted from those expected by the weak-lensing mass and those for the other two clusters show no significant deviations, which is consistent with predictions of numerical simulations

A blood RNA signature for tuberculosis disease risk: a prospective cohort study.

BACKGROUND: Identification of blood biomarkers that prospectively predict progression of Mycobacterium tuberculosis infection to tuberculosis disease might lead to interventions that combat the tuberculosis epidemic. We aimed to assess whether global gene expression measured in whole blood of healthy people allowed identification of prospective signatures of risk of active tuberculosis disease. METHODS: In this prospective cohort study, we followed up healthy, South African adolescents aged 12-18 years from the adolescent cohort study (ACS) who were infected with M tuberculosis for 2 years. We collected blood samples from study participants every 6 months and monitored the adolescents for progression to tuberculosis disease. A prospective signature of risk was derived from whole blood RNA sequencing data by comparing participants who developed active tuberculosis disease (progressors) with those who remained healthy (matched controls). After adaptation to multiplex quantitative real-time PCR (qRT-PCR), the signature was used to predict tuberculosis disease in untouched adolescent samples and in samples from independent cohorts of South African and Gambian adult progressors and controls. Participants of the independent cohorts were household contacts of adults with active pulmonary tuberculosis disease. FINDINGS: Between July 6, 2005, and April 23, 2007, we enrolled 6363 participants from the ACS study and 4466 from independent South African and Gambian cohorts. 46 progressors and 107 matched controls were identified in the ACS cohort. A 16 gene signature of risk was identified. The signature predicted tuberculosis progression with a sensitivity of 66·1% (95% CI 63·2-68·9) and a specificity of 80·6% (79·2-82·0) in the 12 months preceding tuberculosis diagnosis. The risk signature was validated in an untouched group of adolescents (p=0·018 for RNA sequencing and p=0·0095 for qRT-PCR) and in the independent South African and Gambian cohorts (p values <0·0001 by qRT-PCR) with a sensitivity of 53·7% (42·6-64·3) and a specificity of 82·8% (76·7-86) in the 12 months preceding tuberculosis. INTERPRETATION: The whole blood tuberculosis risk signature prospectively identified people at risk of developing active tuberculosis, opening the possibility for targeted intervention to prevent the disease. FUNDING: Bill & Melinda Gates Foundation, the National Institutes of Health, Aeras, the European Union, and the South African Medical Research Council

A Bayesian view of murine seminal cytokine networks

It has long been established that active agents in seminal fluid are key to initiating and coordinating mating-induced immunomodulation. This is in part governed by the actions of a network of cytokine interactions which, to date, remain largely undefined, and whose interspecific evolutionary conservation is unknown. This study applied Bayesian methods to illustrate the interrelationships between seminal profiles of interleukin (IL)-1alpha, IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12 (p70), IL-13, IL-17, eotaxin, granulocyte-colony stimulating factor (G-CSF), granulocyte macrophage-colony stimulating factor (GM-CSF), interferon (IFN)-gamma, keratinocyte-derived chemokine (KC), monocyte chemoattractant protein (MCP-1), macrophage inflammatory protein (MIP-1) alpha, MIP-1beta, regulated on activation normal T cell expressed and secreted (RANTES), tumour necrosis factor (TNF)-alpha, leptin, inducible protein (IP)-10 and vascular endothelial growth factor (VEGF) in a rat model. IL-2, IL-9, IL-12 (p70), IL-13, IL-18, eotaxin, IFN-gamma, IP-10, KC, leptin, MCP-1, MIP-1alpha and TNF-alpha were significantly higher in serum, whilst IL-1beta, IL-5, IL-6, IL-10, IL-17, G-CSF and GM-CSF were significantly higher in seminal fluid. When compared to mouse profiles, only G-CSF was present at significantly higher levels in the seminal fluid in both species. Bayesian modelling highlighted key shared features across mouse and rat networks, namely TNF-alpha as the terminal node in both serum and seminal plasma, and MCP-1 as a central coordinator of seminal cytokine networks through the intermediary of KC and RANTES. These findings reveal a marked interspecific conservation of seminal cytokine networks

Four-Gene Pan-African Blood Signature Predicts Progression to Tuberculosis.

Rationale: Contacts of patients with tuberculosis (TB) constitute an important target population for preventive measures because they are at high risk of infection with Mycobacterium tuberculosis and progression to disease.Objectives: We investigated biosignatures with predictive ability for incident TB.Methods: In a case-control study nested within the Grand Challenges 6-74 longitudinal HIV-negative African cohort of exposed household contacts, we employed RNA sequencing, PCR, and the pair ratio algorithm in a training/test set approach. Overall, 79 progressors who developed TB between 3 and 24 months after diagnosis of index case and 328 matched nonprogressors who remained healthy during 24 months of follow-up were investigated.Measurements and Main Results: A four-transcript signature derived from samples in a South African and Gambian training set predicted progression up to two years before onset of disease in blinded test set samples from South Africa, the Gambia, and Ethiopia with little population-associated variability, and it was also validated in an external cohort of South African adolescents with latent M. tuberculosis infection. By contrast, published diagnostic or prognostic TB signatures were predicted in samples from some but not all three countries, indicating site-specific variability. Post hoc meta-analysis identified a single gene pair, C1QC/TRAV27 (complement C1q C-chain / T-cell receptor-α variable gene 27) that would consistently predict TB progression in household contacts from multiple African sites but not in infected adolescents without known recent exposure events.Conclusions: Collectively, we developed a simple whole blood-based PCR test to predict TB in recently exposed household contacts from diverse African populations. This test has potential for implementation in national TB contact investigation programs

Measurement of serum hepcidin-25 levels as a potential test for diagnosing hemochromatosis and related disorders

石川県立中央病院金沢大学医薬保健研究域医学系Iron overload syndromes include a wide spectrum of genetic and acquired conditions. Recent studies suggest suppressed hepcidin synthesis in the liver to be the molecular basis of hemochromatosis. However, a liver with acquired iron overload synthesizes an adequate amount of hepcidin. Thus, hepcidin could function as a biochemical marker for differential diagnosis of iron overload syndromes. Methods We measured serum iron parameters and hepcidin- 25 levels followed by sequencing HFE, HJV, HAMP, TFR2, and SLC40A1 genes in 13 Japanese patients with iron overload syndromes. In addition, we performed direct measurement of serum hepcidin-25 levels using liquid chromatography-tandem mass spectrometry in 3 Japanese patients with aceruloplasminemia and 4 Italians with HFE hemochromatosis. Results One patient with HJV hemochromatosis, 2 with TFR2 hemochromatosis, and 3 with ferroportin disease were found among the 13 Japanese patients. The remaining 7 Japanese patients showed no evidence for genetic basis of iron overload syndrome. As far as the serum hepcidin-25 was concerned, seven patients with hemochromatosis and 3 with aceruloplasminemia showed markedly decreased serum hepcidin-25 levels. In contrast, 3 patients with ferroportin disease and 7 with secondary iron overload syndromes showed serum hepcidin levels parallel to their hyperferritinemia. Patients with iron overload syndromes were divided into 2 phenotypes presenting as low and high hepcidinemia. These were then associated with their genotypes. Conclusion Determining serum hepcidin-25 levels may aid differential diagnosis of iron overload syndromes prior to genetic analysis. © Springer 2010

MUC1 Limits Helicobacter pylori Infection both by Steric Hindrance and by Acting as a Releasable Decoy

The bacterium Helicobacter pylori can cause peptic ulcer disease, gastric adenocarcinoma and MALT lymphoma. The cell-surface mucin MUC1 is a large glycoprotein which is highly expressed on the mucosal surface and limits the density of H. pylori in a murine infection model. We now demonstrate that by using the BabA and SabA adhesins, H. pylori bind MUC1 isolated from human gastric cells and MUC1 shed into gastric juice. Both H. pylori carrying these adhesins, and beads coated with MUC1 antibodies, induced shedding of MUC1 from MKN7 human gastric epithelial cells, and shed MUC1 was found bound to H. pylori. Shedding of MUC1 from non-infected cells was not mediated by the known MUC1 sheddases ADAM17 and MMP-14. However, knockdown of MMP-14 partially affected MUC1 release early in infection, whereas ADAM17 had no effect. Thus, it is likely that shedding is mediated both by proteases and by disassociation of the non-covalent interaction between the α- and β-subunits. H. pylori bound more readily to MUC1 depleted cells even when the bacteria lacked the BabA and SabA adhesins, showing that MUC1 inhibits attachment even when bacteria cannot bind to the mucin. Bacteria lacking both the BabA and SabA adhesins caused less apoptosis in MKN7 cells than wild-type bacteria, having a greater effect than deletion of the CagA pathogenicity gene. Deficiency of MUC1/Muc1 resulted in increased epithelial cell apoptosis, both in MKN7 cells in vitro, and in H. pylori infected mice. Thus, MUC1 protects the epithelium from non-MUC1 binding bacteria by inhibiting adhesion to the cell surface by steric hindrance, and from MUC1-binding bacteria by acting as a releasable decoy