The DNA Glycosylases Ogg1 and Nth1 Do Not Contribute to Ig Class Switching in Activated Mouse Splenic B Cells

During activation of B cells to undergo class switching, B cell metabolism is increased, and levels of reactive oxygen species (ROS) are increased. ROS can oxidize DNA bases resulting in substrates for the DNA glycosylases Ogg1 and Nth1. Ogg1 and Nth1 excise oxidized bases, and nick the resulting abasic sites, forming single-strand DNA breaks (SSBs) as intermediates during the repair process. In this study, we asked whether splenic B cells from mice deficient in these two enzymes would show altered class switching and decreased DNA breaks in comparison with wild-type mice. As the c-myc gene frequently recombines with the IgH S region in B cells induced to undergo class switching, we also analyzed the effect of deletion of these two glycosylases on DSBs in the c-myc gene. We did not detect a reduction in S region or c-myc DSBs or in class switching in splenic B cells from Ogg1- or Nth1-deficient mice or from mice deficient in both enzymes

Giant hydronephrosis mimicking progressive malignancy

BACKGROUND: Cases of giant hydronephroses are rare and usually contain no more than 1–2 litres of fluid in the collecting system. We report a remarkable case of giant hydronephrosis mimicking a progressive malignant abdominal tumour. CASE PRESENTATION: A 78-year-old cachectic woman presented with an enormous abdominal tumour, which, according to the patient, had slowly increased in diameter. Medical history was unremarkable except for a hysterectomy >30 years before. A CT scan revealed a giant cystic tumour filling almost the entire abdominal cavity. It was analysed by two independent radiologists who suspected a tumour originating from the right kidney and additionally a cystic ovarian neoplasm. Subsequently, a diagnostic and therapeutic laparotomy was performed: the tumour presented as a cystic, 35 × 30 × 25 cm expansive structure adhesive to adjacent organs without definite signs of invasive growth. The right renal hilar vessels could finally be identified at its basis. After extirpation another tumourous structure emerged in the pelvis originating from the genital organs and was also resected. The histopathological examination revealed a >15 kg hydronephrotic right kidney, lacking hardly any residual renal cortex parenchyma. The second specimen was identified as an ovary with regressive changes and a large partially calcified cyst. There was no evidence of malignant growth. CONCLUSION: Although both clinical symptoms and the enormous size of the tumour indicated malignant growth, it turned out to be a giant hydronephrosis. Presumably, a chronic obstruction of the distal ureter had caused this extraordinary hydronephrosis. As demonstrated in our case, an accurate diagnosis of giant hydronephrosis remains challenging due to the atrophy of the renal parenchyma associated with chronic obstruction. Therefore, any abdominal cystic mass even in the absence of other evident pathologies should include the differential diagnosis of a possible hydronephrosis. Diagnostic accuracy might be increased by a combination of endourological techniques such as retrograde pyelography and modern imaging modalities

The making of a mammalian peroxisome, version 2.0: mitochondria get into the mix

This is the author accepted manuscript. The final version is available from Nature Publishing Group via the DOI in this record.A recent report from the laboratory of Heidi McBride (McGill University) presents a role for mitochondria in the de novo biogenesis of peroxisomes in mammalian cells (1). Peroxisomes are essential organelles responsible for a wide variety of biochemical functions, from the generation of bile, to plasmalogen synthesis, reduction of peroxides, and the oxidation of very long chain fatty acids (2). Like mitochondria, peroxisomes proliferate primarily through growth and division of pre-existing peroxisomes (3-6). However, unlike mitochondria, peroxisomes do not fuse (5,7); further, and perhaps most importantly, they can also be born de novo, a process thought to occur through the generation of pre-peroxisomal vesicles that originate from the endoplasmic reticulum (reviewed in (8,9). De novo peroxisome biogenesis has been extensively studies in yeast, with a major focus on the role of the ER in this process. Comprehensive studies in mammalian cells are, however, scarce (5,10-12). By exploiting patient cells lacking mature peroxisomes, Sugiura et al. (1) now assign a role to ER and mitochondria in de novo mammalian peroxisome biogenesis by showing that the formation of immature preperoxisomes occurs through the fusion of Pex3- / Pex14-containing mitochondriaderived vesicles with Pex16-containing ER-derived vesicles

Metabolite Profiling Uncovers Plasmid-Induced Cobalt Limitation under Methylotrophic Growth Conditions

BACKGROUND:The introduction and maintenance of plasmids in cells is often associated with a reduction of growth rate. The reason for this growth reduction is unclear in many cases. METHODOLOGY/PRINCIPAL FINDINGS:We observed a surprisingly large reduction in growth rate of about 50% of Methylobacterium extorquens AM1 during methylotrophic growth in the presence of a plasmid, pCM80 expressing the tetA gene, relative to the wild-type. A less pronounced growth delay during growth under non-methylotrophic growth conditions was observed; this suggested an inhibition of one-carbon metabolism rather than a general growth inhibition or metabolic burden. Metabolome analyses revealed an increase in pool sizes of ethylmalonyl-CoA and methylmalonyl-CoA of more than 6- and 35-fold, respectively, relative to wild type, suggesting a strongly reduced conversion of these central intermediates, which are essential for glyoxylate regeneration in this model methylotroph. Similar results were found for M. extorquens AM1 pCM160 which confers kanamycin resistance. These intermediates of the ethylmalonyl-CoA pathway have in common their conversion by coenzyme B(12)-dependent mutases, which have cobalt as a central ligand. The one-carbon metabolism-related growth delay was restored by providing higher cobalt concentrations, by heterologous expression of isocitrate lyase as an alternative path for glyoxylate regeneration, or by identification and overproduction of proteins involved in cobalt import. CONCLUSIONS/SIGNIFICANCE:This study demonstrates that the introduction of the plasmids leads to an apparent inhibition of the cobalt-dependent enzymes of the ethylmalonyl-CoA pathway. Possible explanations are presented and point to a limited cobalt concentration in the cell as a consequence of the antibiotic stress

Influence of bevacizumab, sunitinib and sorafenib as single agents or in combination on the inhibitory effects of VEGF on human dendritic cell differentiation from monocytes

Vascular endothelial growth factor (VEGF) inhibits differentiation and maturation of dendritic cells (DC), suggesting a potential immunosuppressive role for this proangiogenic factor. Bevacizumab, sorafenib and sunitinib target VEGF-mediated angiogenesis and are active against several types of cancer, but their effects on the immune system are poorly understood. In this study, VEGF and supernatants of renal carcinoma cell lines cultured under hypoxia were found to alter the differentiation of human monocytes to DC. Resulting DC showed impaired activity, as assessed by the alloreactive mixed T-lymphocyte reaction. Bevacizumab and sorafenib, but not sunitinib, reversed the inhibitory effects of VEGF, but not of those mediated by tumour supernatants. Dendritic cells matured under the influence of VEGF expressed less human leukocyte antigen-DR (HLA-DR) and CD86, and this effect was restored by bevacizumab and sorafenib. Finally, tumour-cell supernatants decreased interleukin-12 (IL-12) production by mature DC, and such inhibition was not restored by any of the tested drugs, delivered either as single agents or in combination. The deleterious effects of tumour-cell supernatants were mainly mediated by thermostable molecules distinct from VEGF. These results indicate that inhibition of the differentiation of monocytes to DC is a multifactorial effect, and that they support the development of combinations of angiogenesis inhibitors with immunological modulators

Mining recent brain proteomic databases for ion channel phosphosite nuggets

Voltage-gated ion channels underlie electrical activity of neurons and are dynamically regulated by diverse cell signaling pathways that alter their phosphorylation state. Recent global mass spectrometric–based analyses of the mouse brain phosphoproteome have yielded a treasure trove of new data as to the extent and nature of phosphorylation of numerous ion channel principal or α subunits in mammalian brain. Here we compile and review data on 347 phosphorylation sites (261 unique) on 42 different voltage-gated ion channel α subunits that were identified in these recent studies. Researchers in the ion channel field can now begin to explore the role of these novel in vivo phosphorylation sites in the dynamic regulation of the localization, activity, and expression of brain ion channels through multisite phosphorylation of their principal subunits

Antibodies for Assessing Circadian Clock Proteins in the Rodent Suprachiasmatic Nucleus

Research on the mechanisms underlying circadian rhythmicity and the response of brain and body clocks to environmental and physiological challenges requires assessing levels of circadian clock proteins. Too often, however, it is difficult to acquire antibodies that specifically and reliably label these proteins. Many of these antibodies also lack appropriate validation. The goal of this project was to generate and characterize antibodies against several circadian clock proteins. We examined mice and hamsters at peak and trough times of clock protein expression in the suprachiasmatic nucleus (SCN). In addition, we confirmed specificity by testing the antibodies on mice with targeted disruption of the relevant genes. Our results identify antibodies against PER1, PER2, BMAL1 and CLOCK that are useful for assessing circadian clock proteins in the SCN by immunocytochemistry

LYL1 Degradation by the Proteasome Is Directed by a N-Terminal PEST Rich Site in a Phosphorylation-Independent Manner

Background: The Lymphoblastic leukemia 1 (LYL1) gene is a proto-oncogenic transcription factor found upregulated in patients with T-cell acute lymphoblastic leukemia (T-cell ALL). Initially, the upregulation was described to be as a result of a translocation. However, further studies revealed that transcriptional upregulation of LYL1could also occur without translocations. In addition, post-translational mechanisms, such as protein degradation could influence LYL1 expression as well. Methodology/Principal Findings: In this study, we considered possible post-translational regulation of Lyl1, and investigated fundamental mechanisms governing LYL1 degradation in cell-based culture assays. We identify a PEST sequence motif located in the N-terminus of LYL1, which determines the efficiency of LYL1 degradation by the proteasome. The absence of the PEST degradation site leads to accumulation or upregulation of LYL1. We also show that LYL1 is phosphorylated by MAPK at S36, and determined that proteasomal degradation of LYL1 occurs in a phosphorylationindependent manner. Conclusions/Significance: Understanding LYL1 degradation is a step forward not only towards deciphering the normal function and regulation of LYL1, but could suggest post-translational mechanisms for upregulation of LYL1 that ma

TRY plant trait database - enhanced coverage and open access

Plant traits-the morphological, anatomical, physiological, biochemical and phenological characteristics of plants-determine how plants respond to environmental factors, affect other trophic levels, and influence ecosystem properties and their benefits and detriments to people. Plant trait data thus represent the basis for a vast area of research spanning from evolutionary biology, community and functional ecology, to biodiversity conservation, ecosystem and landscape management, restoration, biogeography and earth system modelling. Since its foundation in 2007, the TRY database of plant traits has grown continuously. It now provides unprecedented data coverage under an open access data policy and is the main plant trait database used by the research community worldwide. Increasingly, the TRY database also supports new frontiers of trait-based plant research, including the identification of data gaps and the subsequent mobilization or measurement of new data. To support this development, in this article we evaluate the extent of the trait data compiled in TRY and analyse emerging patterns of data coverage and representativeness. Best species coverage is achieved for categorical traits-almost complete coverage for 'plant growth form'. However, most traits relevant for ecology and vegetation modelling are characterized by continuous intraspecific variation and trait-environmental relationships. These traits have to be measured on individual plants in their respective environment. Despite unprecedented data coverage, we observe a humbling lack of completeness and representativeness of these continuous traits in many aspects. We, therefore, conclude that reducing data gaps and biases in the TRY database remains a key challenge and requires a coordinated approach to data mobilization and trait measurements. This can only be achieved in collaboration with other initiatives