PTPα regulates integrin-stimulated FAK autophosphorylation and cytoskeletal rearrangement in cell spreading and migration

We investigated the molecular and cellular actions of receptor protein tyrosine phosphatase (PTP) α in integrin signaling using immortalized fibroblasts derived from wild-type and PTPα-deficient mouse embryos. Defects in PTPα−/− migration in a wound healing assay were associated with altered cell shape and focal adhesion kinase (FAK) phosphorylation. The reduced haptotaxis to fibronectin (FN) of PTPα−/− cells was increased by expression of active (but not inactive) PTPα. Integrin-mediated formation of src–FAK and fyn–FAK complexes was reduced or abolished in PTPα−/− cells on FN, concomitant with markedly reduced phosphorylation of FAK at Tyr397. Reintroduction of active (but not inactive) PTPα restored FAK Tyr-397 phosphorylation. FN-induced cytoskeletal rearrangement was retarded in PTPα−/− cells, with delayed filamentous actin stress fiber assembly and focal adhesion formation. This mimicked the effects of treating wild-type fibroblasts with the src family protein tyrosine kinase (Src-PTK) inhibitor PP2. These results, together with the reduced src/fyn tyrosine kinase activity in PTPα−/− fibroblasts (Ponniah et al., 1999; Su et al., 1999), suggest that PTPα functions in integrin signaling and cell migration as an Src-PTK activator. Our paper establishes that PTPα is required for early integrin-proximal events, acting upstream of FAK to affect the timely and efficient phosphorylation of FAK Tyr-397

Characterization of Endothelial Cells Associated with Hematopoietic Niche Formation in Humans Identifies IL-33 As an Anabolic Factor

Bone marrow formation requires an orchestrated interplay between osteogenesis, angiogenesis, and hematopoiesis that is thought to be mediated by endothelial cells. The nature of the endothelial cells and the molecular mechanisms underlying these events remain unclear in humans. Here, we identify a subset of endoglin-expressing endothelial cells enriched in human bone marrow during fetal ontogeny and upon regeneration after chemotherapeutic injury. Comprehensive transcriptional characterization by massive parallel RNA sequencing of these cells reveals a phenotypic and molecular similarity to murine type H endothelium and activation of angiocrine factors implicated in hematopoiesis, osteogenesis, and angiogenesis. Interleukin-33 (IL-33) was significantly overexpressed in these endothelial cells and promoted the expansion of distinct subsets of h

The Genes Coding for the Conversion of Carbazole to Catechol Are Flanked by IS6100 Elements in Sphingomonas sp. Strain XLDN2-5

BACKGROUND: Carbazole is a recalcitrant compound with a dioxin-like structure and possesses mutagenic and toxic activities. Bacteria respond to a xenobiotic by recruiting exogenous genes to establish a pathway to degrade the xenobiotic, which is necessary for their adaptation and survival. Usually, this process is mediated by mobile genetic elements such as plasmids, transposons, and insertion sequences. FINDINGS: The genes encoding the enzymes responsible for the degradation of carbazole to catechol via anthranilate were cloned, sequenced, and characterized from a carbazole-degrading Sphingomonas sp. strain XLDN2-5. The car gene cluster (carRAaBaBbCAc) and fdr gene were accompanied on both sides by two copies of IS6100 elements, and organized as IS6100::ISSsp1-ORF1-carRAaBaBbCAc-ORF8-IS6100-fdr-IS6100. Carbazole was converted by carbazole 1,9a-dioxygenase (CARDO, CarAaAcFdr), meta-cleavage enzyme (CarBaBb), and hydrolase (CarC) to anthranilate and 2-hydroxypenta-2,4-dienoate. The fdr gene encoded a novel ferredoxin reductase whose absence resulted in lower transformation activity of carbazole by CarAa and CarAc. The ant gene cluster (antRAcAdAbAa) which was involved in the conversion of anthranilate to catechol was also sandwiched between two IS6100 elements as IS6100-antRAcAdAbAa-IS6100. Anthranilate 1,2-dioxygenase (ANTDO) was composed of a reductase (AntAa), a ferredoxin (AntAb), and a two-subunit terminal oxygenase (AntAcAd). Reverse transcription-PCR results suggested that carAaBaBbCAc gene cluster, fdr, and antRAcAdAbAa gene cluster were induced when strain XLDN2-5 was exposed to carbazole. Expression of both CARDO and ANTDO in Escherichia coli required the presence of the natural reductases for full enzymatic activity. CONCLUSIONS/SIGNIFICANCE: We predict that IS6100 might play an important role in the establishment of carbazole-degrading pathway, which endows the host to adapt to novel compounds in the environment. The organization of the car and ant genes in strain XLDN2-5 was unique, which showed strong evolutionary trail of gene recruitment mediated by IS6100 and presented a remarkable example of rearrangements and pathway establishments

The Threonine Protease Activity of Testes-Specific Protease 50 (TSP50) Is Essential for Its Function in Cell Proliferation

Background: Testes-specific protease 50 (TSP50), a newly discovered threonine enzyme, has similar amino acid sequences and enzymatic structures to those of many serine proteases. It may be an oncogene. TSP50 is up-regulated in breast cancer epithelial cells, and ectopic expression of TSP50 in TSP50-deficient Chinese hamster ovary (CHO) cells has been found to promote cell proliferation. However, the mechanisms by which TSP50 exerts its growth-promoting effects are not yet fully understood. Methodology/Principal Findings: To delineate whether the threonine protease activity of TSP50 is essential to its function in cell proliferation, we constructed and characterized a mutant TSP50, called TSP50 T310A, which was identified as a protease-dead mutant of TSP50. By a series of proliferation analyses, colony formation assays and apoptosis analyses, we showed that T310A mutation significantly depresses TSP50-induced cell proliferation in vitro. Next, the CHO stable cell line expressing either wild-type or T310A mutant TSP50 was injected subcutaneously into nude mice. We found that the T310A mutation could abolish the tumorigenicity of TSP50 in vivo. A mechanism investigation revealed that the T310A mutation prevented interaction between TSP50 and the NF-kBIkBa complex, which is necessary for TSP50 to perform its function in cell proliferation. Conclusion: Our data highlight the importance of threonine 310, the most critical protease catalytic site in TSP50, to TSP50induce

Measurement of the inclusive and dijet cross-sections of b-jets in pp collisions at sqrt(s) = 7 TeV with the ATLAS detector

The inclusive and dijet production cross-sections have been measured for jets containing b-hadrons (b-jets) in proton-proton collisions at a centre-of-mass energy of sqrt(s) = 7 TeV, using the ATLAS detector at the LHC. The measurements use data corresponding to an integrated luminosity of 34 pb^-1. The b-jets are identified using either a lifetime-based method, where secondary decay vertices of b-hadrons in jets are reconstructed using information from the tracking detectors, or a muon-based method where the presence of a muon is used to identify semileptonic decays of b-hadrons inside jets. The inclusive b-jet cross-section is measured as a function of transverse momentum in the range 20 < pT < 400 GeV and rapidity in the range |y| < 2.1. The bbbar-dijet cross-section is measured as a function of the dijet invariant mass in the range 110 < m_jj < 760 GeV, the azimuthal angle difference between the two jets and the angular variable chi in two dijet mass regions. The results are compared with next-to-leading-order QCD predictions. Good agreement is observed between the measured cross-sections and the predictions obtained using POWHEG + Pythia. MC@NLO + Herwig shows good agreement with the measured bbbar-dijet cross-section. However, it does not reproduce the measured inclusive cross-section well, particularly for central b-jets with large transverse momenta.Comment: 10 pages plus author list (21 pages total), 8 figures, 1 table, final version published in European Physical Journal

Jet energy measurement with the ATLAS detector in proton-proton collisions at root s=7 TeV

The jet energy scale and its systematic uncertainty are determined for jets measured with the ATLAS detector at the LHC in proton-proton collision data at a centre-of-mass energy of √s = 7TeV corresponding to an integrated luminosity of 38 pb-1. Jets are reconstructed with the anti-kt algorithm with distance parameters R=0. 4 or R=0. 6. Jet energy and angle corrections are determined from Monte Carlo simulations to calibrate jets with transverse momenta pT≥20 GeV and pseudorapidities {pipe}η{pipe}<4. 5. The jet energy systematic uncertainty is estimated using the single isolated hadron response measured in situ and in test-beams, exploiting the transverse momentum balance between central and forward jets in events with dijet topologies and studying systematic variations in Monte Carlo simulations. The jet energy uncertainty is less than 2. 5 % in the central calorimeter region ({pipe}η{pipe}<0. 8) for jets with 60≤pT<800 GeV, and is maximally 14 % for pT<30 GeV in the most forward region 3. 2≤{pipe}η{pipe}<4. 5. The jet energy is validated for jet transverse momenta up to 1 TeV to the level of a few percent using several in situ techniques by comparing a well-known reference such as the recoiling photon pT, the sum of the transverse momenta of tracks associated to the jet, or a system of low-pT jets recoiling against a high-pT jet. More sophisticated jet calibration schemes are presented based on calorimeter cell energy density weighting or hadronic properties of jets, aiming for an improved jet energy resolution and a reduced flavour dependence of the jet response. The systematic uncertainty of the jet energy determined from a combination of in situ techniques is consistent with the one derived from single hadron response measurements over a wide kinematic range. The nominal corrections and uncertainties are derived for isolated jets in an inclusive sample of high-pT jets. Special cases such as event topologies with close-by jets, or selections of samples with an enhanced content of jets originating from light quarks, heavy quarks or gluons are also discussed and the corresponding uncertainties are determined. © 2013 CERN for the benefit of the ATLAS collaboration

Induction of Apoptosis Coupled to Endoplasmic Reticulum Stress in Human Prostate Cancer Cells by n-butylidenephthalide

BACKGROUND: N-butylidenephthalide (BP) exhibits antitumor effect in a variety of cancer cell lines. The objective of this study was to obtain additional insights into the mechanisms involved in BP induced cell death in human prostate cancer cells. METHODS/PRINCIPAL FINDINGS: Two human prostate cancer cell lines, PC-3 and LNCaP, were treated with BP, and subsequently evaluated for their viability and cell cycle profiles. BP caused cell cycle arrest and cell death in both cell lines. The G0/G1 phase arrest was correlated with increase levels of CDK inhibitors (p16, p21 and p27) and decrease of the checkpoint proteins. To determine the mechanisms of BP-induced growth arrest and cell death in prostate cancer cell lines, we performed a microarray study to identify alterations in gene expression induced by BP in the LNCaP cells. Several BP-induced genes, including the GADD153/CHOP, an endoplasmic reticulum stress (ER stress)-regulated gene, were identified. BP-induced ER stress was evidenced by increased expression of the downstream molecules GRP78/BiP, IRE1-α and GADD153/CHOP in both cell lines. Blockage of IRE1-α or GADD153/CHOP expression by siRNA significantly reduced BP-induced cell death in LNCaP cells. Furthermore, blockage of JNK1/2 signaling by JNK siRNA resulted in decreased expression of IRE1-α and GADD153/CHOP genes, implicating that BP-induced ER stress may be elicited via JNK1/2 signaling in prostate cancer cells. BP also suppressed LNCaP xenograft tumor growth in NOD-SCID mice. It caused 68% reduction in tumor volume after 18 days of treatment. CONCLUSIONS: Our results suggest that BP can cause G0/G1 phase arrest in prostate cancer cells and its cytotoxicity is mediated by ER stress induction. Thus, BP may serve as an anticancer agent by inducing ER stress in prostate cancer

Activation of NF-κB driven inflammatory programs in mesenchymal elements attenuates hematopoiesis in low-risk myelodysplastic syndromes

Activation of NF-κB signaling in mesenchymal cells is common in LR-MDS.Activation of NF-κB in mesenchymal cells leads to transcriptional overexpression of inflammatory factors including negative regulators of hematopoiesis.Activation of NF-κB attenuates HSPC numbers and function ex vivo