The Rewiring of Ubiquitination Targets in a Pathogenic Yeast Promotes Metabolic Flexibility, Host Colonization and Virulence

Funding: This work was funded by the European Research Council [http://erc.europa.eu/], AJPB (STRIFE Advanced Grant; C-2009-AdG-249793). The work was also supported by: the Wellcome Trust [www.wellcome.ac.uk], AJPB (080088, 097377); the UK Biotechnology and Biological Research Council [www.bbsrc.ac.uk], AJPB (BB/F00513X/1, BB/K017365/1); the CNPq-Brazil [http://cnpq.br], GMA (Science without Borders fellowship 202976/2014-9); and the National Centre for the Replacement, Refinement and Reduction of Animals in Research [www.nc3rs.org.uk], DMM (NC/K000306/1). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Acknowledgments We thank Dr. Elizabeth Johnson (Mycology Reference Laboratory, Bristol) for providing strains, and the Aberdeen Proteomics facility for the biotyping of S. cerevisiae clinical isolates, and to Euroscarf for providing S. cerevisiae strains and plasmids. We are grateful to our Microscopy Facility in the Institute of Medical Sciences for their expert help with the electron microscopy, and to our friends in the Aberdeen Fungal Group for insightful discussions.Peer reviewedPublisher PD

LRRK2 at the interface of autophagosomes, endosomes and lysosomes

Over the past 20 years, substantial progress has been made in identifying the underlying genetics of Parkinson’s disease (PD). Of the known genes, LRRK2 is a major genetic contributor to PD. However, the exact function of LRRK2 remains to be elucidated. In this review, we discuss how familial forms of PD have led us to hypothesize that alterations in endomembrane trafficking play a role in the pathobiology of PD. We will discuss the major observations that have been made to elucidate the role of LRRK2 in particular, including LRRK2 animal models and high-throughput proteomics approaches. Taken together, these studies strongly support a role of LRRK2 in vesicular dynamics. We also propose that targeting these pathways may not only be beneficial for developing therapeutics for LRRK2-driven PD, but also for other familial and sporadic cases

Cell-based screen for altered nuclear phenotypes reveals senescence progression in polyploid cells after Aurora kinase B inhibition.

Cellular senescence is a widespread stress response and is widely considered to be an alternative cancer therapeutic goal. Unlike apoptosis, senescence is composed of a diverse set of subphenotypes, depending on which of its associated effector programs are engaged. Here we establish a simple and sensitive cell-based prosenescence screen with detailed validation assays. We characterize the screen using a focused tool compound kinase inhibitor library. We identify a series of compounds that induce different types of senescence, including a unique phenotype associated with irregularly shaped nuclei and the progressive accumulation of G1 tetraploidy in human diploid fibroblasts. Downstream analyses show that all of the compounds that induce tetraploid senescence inhibit Aurora kinase B (AURKB). AURKB is the catalytic component of the chromosome passenger complex, which is involved in correct chromosome alignment and segregation, the spindle assembly checkpoint, and cytokinesis. Although aberrant mitosis and senescence have been linked, a specific characterization of AURKB in the context of senescence is still required. This proof-of-principle study suggests that our protocol is capable of amplifying tetraploid senescence, which can be observed in only a small population of oncogenic RAS-induced senescence, and provides additional justification for AURKB as a cancer therapeutic target.This work was supported by the University of Cambridge, Cancer Research UK, Hutchison Whampoa; Cancer Research UK grants A6691 and A9892 (M.N., N.K., C.J.T., D.C.B., C.J.C., L.S.G, and M.S.); a fellowship from the Uehara Memorial Foundation (M.S.).This is the author accepted manuscript. The final version is available from the American Society for Cell Biology via http://dx.doi.org/10.1091/mbc.E15-01-000

Cerebral palsy aetiologic diagnosis

A retrospective study was carried on 35 cases with Cerebral Palsy, obtained at random, attended at the Hospital of Clinics of School Medicine of São Paulo University, Campus Ribeirão Preto, from 1982 to 1998, with the scope to verify the aetiologic factors. Among 25 patients with identified aetiology, the Congenital infections (36%) and hypoxic-schemic cerebral lesions (28%) were the major causes. These results could be used for regional strategies of deficiencies prevention. Comments about the cases without aetiology identified points to the need to consider the inherited cerebral palsies and the differential diagnosis with degenerative diseases.Em estudo retrospectivo foram analisados 35 prontuários, obtidos ao acaso, de crianças com diagnóstico de Paralisia Cerebral (PC), atendidas no Hospital das Clínicas de Ribeirão Preto, entre 1982 e 1998, com o objetivo de verificar fatores etiológicos. As infecções congênitas (36%) e agressão hipóxico-isquêmica (28%) foram os mais freqüentes fatores determinantes de PC entre 25 casos com etiologia definida. Estes achados podem ser utilizados para estratégias regionais de prevenção de deficiências. Para os casos sem etiologia definida, lembram-se as formas herdadas de paralisia cerebral e o diagnóstico diferencial com doenças degenerativas

Tuberculosis contact investigation with a new, specific blood test in a low-incidence population containing a high proportion of BCG-vaccinated persons

BACKGROUND: BCG-vaccination can confound tuberculin skin test (TST) reactions in the diagnosis of latent tuberculosis infection. METHODS: We compared the TST with a Mycobacterium tuberculosis specific whole blood interferon-gamma assay (QuantiFERON(®)-TB-Gold In Tube; QFT-G) during ongoing investigations among close contacts of sputum smear positive source cases in Hamburg, Germany. RESULTS: During a 6-month period, 309 contacts (mean age 28.5 ± 10.5 years) from a total of 15 source cases underwent both TST and QFT-G testing. Of those, 157 (50.8%) had received BCG vaccination and 84 (27.2%) had migrated to Germany from a total of 25 different high prevalence countries (i.e. >20 cases/100,000). For the TST, the positive response rate was 44.3% (137/309), whilst only 31 (10%) showed a positive QFT-G result. The overall agreement between the TST and the QFT-G was low (κ = 0.2, with 95% CI 0.14.-0.23), and positive TST reactions were closely associated with prior BCG vaccination (OR 24.7; 95% CI 11.7–52.5). In contrast, there was good agreement between TST and QFT-G in non-vaccinated persons (κ = 0.58, with 95% CI 0.4–0.68), increasing to 0.68 (95% CI 0.46–0.81), if a 10-mm cut off for the TST was used instead of the standard 5 mm recommended in Germany. CONCLUSION: The QFT-G assay was unaffected by BCG vaccination status, unlike the TST. In close contacts who were BCG-vaccinated, the QFT-G assay appeared to be a more specific indicator of latent tuberculosis infection than the TST, and similarly sensitive in unvaccinated contacts. In BCG-vaccinated close contacts, measurement of IFN-gamma responses of lymphocytes stimulated with M. tuberculosis-specific antigen should be recommended as a basis for the decision on whether to perform subsequent chest X-ray examinations or to start treatment for latent tuberculosis infection

NF-κB Mediates Tumor Necrosis Factor α-Induced Expression of Optineurin, a Negative Regulator of NF-κB

Optineurin is a ubiquitously expressed multifunctional cytoplasmic protein encoded by OPTN gene. The expression of optineurin is induced by various cytokines. Here we have investigated the molecular mechanisms which regulate optineurin gene expression and the relationship between optineurin and nuclear factor κB (NF-κB). We cloned and characterized human optineurin promoter. Optineurin promoter was activated upon treatment of HeLa and A549 cells with tumor necrosis factor α (TNFα). Mutation of a putative NF-κB-binding site present in the core promoter resulted in loss of basal as well as TNFα-induced activity. Overexpression of p65 subunit of NF-κB activated this promoter through NF-κB site. Oligonucleotides corresponding to this putative NF-κB-binding site showed binding to NF-κB. TNFα-induced optineurin promoter activity was inhibited by expression of inhibitor of NF-κB (IκBα) super-repressor. Blocking of NF-κB activation resulted in inhibition of TNFα-induced optineurin gene expression. Overexpressed optineurin partly inhibited TNFα-induced NF-κB activation in Hela cells. Downregulation of optineurin by shRNA resulted in an increase in TNFα-induced as well as basal NF-κB activity. These results show that optineurin promoter activity and gene expression are regulated by NF-κB pathway in response to TNFα. In addition these results suggest that there is a negative feedback loop in which TNFα-induced NF-κB activity mediates expression of optineurin, which itself functions as a negative regulator of NF-κB