Initial B Cell Activation Induces Metabolic Reprogramming and Mitochondrial Remodeling.

B lymphocytes provide adaptive immunity by generating antigen-specific antibodies and supporting the activation of T cells. Little is known about how global metabolism supports naive B cell activation to enable an effective immune response. By coupling RNA sequencing (RNA-seq) data with glucose isotopomer tracing, we show that stimulated B cells increase programs for oxidative phosphorylation (OXPHOS), the tricarboxylic acid (TCA) cycle, and nucleotide biosynthesis, but not glycolysis. Isotopomer tracing uncovered increases in TCA cycle intermediates with almost no contribution from glucose. Instead, glucose mainly supported the biosynthesis of ribonucleotides. Glucose restriction did not affect B cell functions, yet the inhibition of OXPHOS or glutamine restriction markedly impaired B cell growth and differentiation. Increased OXPHOS prompted studies of mitochondrial dynamics, which revealed extensive mitochondria remodeling during activation. Our results show how B cell metabolism adapts with stimulation and reveals unexpected details for carbon utilization and mitochondrial dynamics at the start of a humoral immune response

Introduction

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/68234/2/10.1177_002188637401000301.pd

Lysosomal dysfunction and impaired autophagy underlie the pathogenesis of amyloidogenic light chain-mediated cardiotoxicity

AL amyloidosis is the consequence of clonal production of amyloidogenic immunoglobulin light chain (LC) proteins, often resulting in a rapidly progressive and fatal amyloid cardiomyopathy. Recent work has found that amyloidogenic LC directly initiate a cardio-toxic response underlying the pathogenesis of the cardiomyopathy; however, the mechanisms that contribute to this proteotoxicity remain unknown. Using human amyloidogenic LC isolated from patients with amyloid cardiomyopathy, we reveal that dysregulation of autophagic flux is critical for mediating amyloidogenic LC proteotoxicity. Restoration of autophagic flux by pharmacological intervention using rapamycin protected against amyloidogenic light chain protein-induced pathologies including contractile dysfunction and cell death at the cellular and organ level and also prolonged survival in an in vivo zebrafish model of amyloid cardiotoxicity. Mechanistically, we identify impaired lysosomal function to be the major cause of defective autophagy and amyloidogenic LC-induced proteotoxicity. Collectively, these findings detail the downstream molecular mechanisms underlying AL amyloid cardiomyopathy and highlight potential targeting of autophagy and lysosomal dysfunction in patients with amyloid cardiomyopathy

Large-scale circulations in a shear-free convective turbulence: Mean-field simulations

It has been previously shown (Phys. Rev. E 66, 066305, 2002) that a non-rotating turbulent convection with nonuniform large-scale flows contributes to the turbulent heat flux. As a result, the turbulent heat flux depends explicitly not only on the gradients of the large-scale temperature, but it also depends on the gradients of the large-scale velocity. This is because the nonuniform large-scale flows produce anisotropic velocity fluctuations which modify the turbulent heat flux. This effect causes an excitation of a convective-wind instability and formation of large-scale semi-organised coherent structures (large-scale convective cells). We perform mean-field numerical simulations of shear-free convection which take into account the modification of the turbulent heat flux by nonuniform large-scale flows. The redistribution of the turbulent heat flux by the nonuniform large-scale motions in turbulent convection plays a crucial role in the formation of the large-scale semi-organised coherent structures. This effect results in a strong reduction of the critical effective Rayleigh number (based on the eddy viscosity and turbulent temperature diffusivity) required for the formation of the large-scale convective cells. The convective-wind instability is excited when the scale separation ratio between the height of the convective layer and the integral turbulence scale is large. The level of the mean kinetic energy at saturation increases with the scale separation ratio. Inside the large-scale convective cells, there are local regions with the positive vertical gradient of the potential temperature which implies that these regions are stably stratified.Comment: 13 pages, 14 figures, revtex4-

Targeting, import, and dimerization of a mammalian mitochondrial ATP binding cassette (ABC) transporter, ABCB10 (ABC-me)

Author Posting. © American Society for Biochemistry and Molecular Biology, 2004. This article is posted here by permission of American Society for Biochemistry and Molecular Biology for personal use, not for redistribution. The definitive version was published in Journal of Biological Chemistry 279 (2004): 42954-42963, doi:10.1074/jbc.M405040200.ATP binding cassette (ABC) transporters are a diverse superfamily of energy-dependent membrane translocases. Although responsible for the majority of transmembrane transport in bacteria, they are relatively uncommon in eukaryotic mitochondria. Organellar trafficking and import, in addition to quaternary structure assembly, of mitochondrial ABC transporters is poorly understood and may offer explanations for the paucity of their diversity. Here we examine these processes in ABCB10 (ABC-me), a mitochondrial inner membrane erythroid transporter involved in heme biosynthesis. We report that ABCB10 possesses an unusually long 105-amino acid mitochondrial targeting presequence (mTP). The central subdomain of the mTP (amino acids (aa) 36–70) is sufficient for mitochondrial import of enhanced green fluorescent protein. The N-terminal subdomain (aa 1–35) of the mTP, although not necessary for the trafficking of ABCB10 to mitochondria, participates in the proper import of the molecule into the inner membrane. We performed a series of amino acid mutations aimed at changing specific properties of the mTP. The mTP requires neither arginine residues nor predictable {alpha}-helices for efficient mitochondrial targeting. Disruption of its hydrophobic character by the mutation L46Q/I47Q, however, greatly diminishes its efficacy. This mutation can be rescued by cryptic downstream (aa 106–715) mitochondrial targeting signals, highlighting the redundancy of this protein's targeting qualities. Mass spectrometry analysis of chemically cross-linked, immunoprecipitated ABCB10 indicates that ABCB10 embedded in the mitochondrial inner membrane homodimerizes and homo-oligomerizes. A deletion mutant of ABCB10 that lacks its mTP efficiently targets to the endoplasmic reticulum. Quaternary structure assembly of ABCB10 in the ER appears to be similar to that in the mitochondria.This work was supported by National Institutes of Health Grants R01HL071629, P41RR001395, and P01HL032262

Thiol modifier effects of diphenyl diselenides: insight from experiment and DFT calculations

A combination of spectroscopic, chromatographic and computational approaches was employed to investigate the reaction of several diselenides of formula (R-PhSe)(2) (R = CH3O, CH3, H, Cl, CF3) with a thiolate nucleophile, leading to the breaking of the selenium-selenium (Se-Se) bond. This process has fundamental importance in biological environments and provides a rationale to analyze the so-called thiol modifier effect of diselenides, which may be exploited in pharmacology and toxicology. Our data suggest that withdrawing substituents favor the reaction, effectively making the reaction energy more negative, but strong electron-withdrawing groups also prompt structural modification on the starting reactant, increasing the reaction barrier. Thus, the nature (electron rich or electron poor) of the diselenides can play an essential role in the reactivity and biological activity of these molecules

Heat transfer to sub-cooled binary solution in horizontal tube

Paper presented at the 5th International Conference on Heat Transfer, Fluid Mechanics and Thermodynamics, South Africa, 1-4 July, 2007.Predicted heat transfer coefficients (HTC) are widely used for calculations of various heat transfer processes. Various empirical and semi-empirical models of non-dimensional groups are used for calculating the heat transfer coefficients. The common accuracy of the predicted heat transfer coefficient is usually about ±25%; however, the accuracy is failing by the inaccurate predictions of the fluid properties. In the present study the heat transfer coefficient of subcooled organic mixture chlorodifluoromethane (R22) - dimethylacetamide (DMAC) were predicted. While the solution’s pressure-temperature-concentration, densitytemperature- concentration and viscosity-temperatureconcentration relations were previously measured in our laboratory and the heat capacity were calculated from the enthalpy of the solution, the only unknown property that was required for predicting the HTC was the mixture thermal conductivity. Various correlations and mixing rules for the mixture thermal conductivity were adopted for predicting the heat transfer coefficient. In order to validate the predicted heat transfer coefficients, an experimental system was designed, built and successfully operated. The heat transfer coefficient was measured experimentally and compared with the predicted one. Based on this comparison the best fitted thermal conductivity for the working fluid (R22-DMAC) is recommended. As a result, the deviations of the predicted heat transfer coefficients, obtained by the well-known Dittus-Boelter equation, from the experimental values were less than 15%.cs201

A novel spontaneous model of epithelial-mesenchymal transition (EMT) using a primary prostate cancer derived cell line demonstrating distinct stem-like characteristics

Cells acquire the invasive and migratory properties necessary for the invasion-metastasis cascade and the establishment of aggressive, metastatic disease by reactivating a latent embryonic programme: epithelial-to-mesenchymal transition (EMT). Herein, we report the development of a new, spontaneous model of EMT which involves four phenotypically distinct clones derived from a primary tumour-derived human prostate cancer cell line (OPCT-1), and its use to explore relationships between EMT and the generation of cancer stem cells (CSCs) in prostate cancer. Expression of epithelial (E-cadherin) and mesenchymal markers (vimentin, fibronectin) revealed that two of the four clones were incapable of spontaneously activating EMT, whereas the others contained large populations of EMT-derived, vimentin-positive cells having spindle-like morphology. One of the two EMT-positive clones exhibited aggressive and stem cell-like characteristics, whereas the other was non-aggressive and showed no stem cell phenotype. One of the two EMT-negative clones exhibited aggressive stem cell-like properties, whereas the other was the least aggressive of all clones. These findings demonstrate the existence of distinct, aggressive CSC-like populations in prostate cancer, but, importantly, that not all cells having a potential for EMT exhibit stem cell-like properties. This unique model can be used to further interrogate the biology of EMT in prostate cancer

Overexpression of multiple oncogenes related to histological grade of astrocytic glioma.

The expression of the c-erbB-1, c-myc, Ha/N-ras and c-fos oncogenes was investigated in 62 astrocytomas of low, intermediate and high grades by immunogold silver histochemistry. Elevated expression of c-erbB-1 was observed in 95%, 48% and 86% of low, intermediate and high grade tumours respectively, c-myc in 5%, 33% and 76% respectively, Ha/N-ras in 0, 43% and 71% respectively and c-fos in 55%, 48% and 52% respectively. Controls included normal brain and tumour sections immunoreacted with pre-immune serum or with antisera absorbed with synthetic peptides. Analysis of co-overexpression revealed that low grade tumours co-overexpressed a maximum of two of these genes, intermediate grade tumours a maximum of three of these genes, while co-overexpression of all four genes was observed in some high grade tumours. Co-overexpression of c-erbB-1 and c-fos was frequently observed in low grade astrocytomas and may be predictive of non-progression. On the other hand, there was a statistically significant increase in the number of tumours overexpressing Ha/N-ras or c-myc with increasing grade of tumour, suggesting that overexpression of these two oncogenes may be indicative of progression

The transcription factor Hey and nuclear lamins specify and maintain cell identity

The inability of differentiated cells to maintain their identity is a hallmark of age-related diseases. We found that the transcription factor Hey supervises the identity of differentiated enterocytes (ECs) in the adult Drosophila midgut. Lineage tracing established that Hey-deficient ECs are unable to maintain their unique nuclear organization and identity. To supervise cell identity, Hey determines the expression of nuclear lamins, switching from a stem-cell lamin configuration to a differentiated lamin configuration. Moreover, continued Hey expression is required to conserve large-scale nuclear organization. During aging, Hey levels decline, and EC identity and gut homeostasis are impaired, including pathological reprograming and compromised gut integrity. These phenotypes are highly similar to those observed upon acute targeting of Hey or perturbation of lamin expression in ECs in young adults. Indeed, aging phenotypes were suppressed by continued expression of Hey in ECs, suggesting that a Hey-lamin network safeguards nuclear organization and differentiated cell identity