Regulation and function of the mitochondrial protease HtrA2/Omi in the control of cell death.

The serine protease HtrA2 is released from mitochondria following apoptotic stimuli. Once in the cytosol, HtrA2 has been implicated in promoting cell death by a caspase-dependent and -independent mechanism. However, mice lacking expression of HtrA2 show no evidence of reduced rates of cell death. On the contrary, loss of HtrA2 causes mitochondrial dysfunction leading to a neurodegenerative disorder with parkinsonian features. This suggests that the protease function of HtrA2 in the mitochondria, and not its pro-apoptotic action in the cytosol, is critical. Mammalian HtrA2 is therefore likely to function in vivo in a manner similar to its bacterial homologues, which are involved in protection against cell stress. The bacterial DegS homologue senses unfolded proteins, activating a proteolytic cascade leading to induction of stress response genes. Transcriptional profiling of wild type and HtrA2 knockout (KO) cells identified the stress-inducible transcription factor CHOP being differentially regulated when mitochondrial stress was triggered. CHOP up-regulation was found in HtrA2 KO mouse brains but not in other tissues. Transcriptional profiling of brain tissue revealed a number of putative ATF4 target genes being up-regulated in HtrA2 KO, among these CHOP. Promoter analysis identified a C/EBP-ATF composite site in the majority of the genes within this signature. Therefore, loss of HtrA2 might impact on nuclear gene expression specifically in brain, subverting normal cellular homeostasis leading to disease. In humans, point mutations in HtrA2 are a susceptibility factor for Parkinson's disease (PD) resulting in partial loss of proteolytic activity. Affinity purification shows that the mitochondrial kinase PINK1 interacts with HtrA2. PINK1 mutations are associated with the PARK6 PD susceptibility locus. HtrA2 is phosphorylated in a PINK1-dependent manner at residues adjacent to positions found mutated in PD patients. Phosphorylation of HtrA2 and thereby modulation of its proteolytic activity seems necessary for the function of HtrA2 in the mitochondria contributing to increased resistance of cells to mitochondrial stress

Inter- and intragrain currents in bulk melt-grown YBaCuO rings

A simple contactless method suitable to discern between the intergrain (circular) current, which flows in the thin superconducting ring, and the intragrain current, which does not cross the weakest link, has been proposed. At first, we show that the intergrain current may directly be estimated from the magnetic flux density $B(\pm z_0)$ measured by the Hall sensor positioned in the special points $\pm z_0$ above/below the ring center. The experimental and the numerical techniques to determine the value $z_0$ are discussed. Being very promising for characterization of a current flowing across the joints in welded YBaCuO rings (its dependencies on the temperature and the external magnetic field as well as the time dissipation), the approach has been applied to study corresponding properties of the intra- and intergrain currents flowing across the $a$-twisted grain boundaries which are frequent in bulk melt-textured YBaCuO samples. We present experimental data related to the flux penetration inside a bore of MT YBaCuO rings both in the non-magnetized, virgin state and during the field reversal. The shielding properties and their dependence on external magnetic fields are also studied. Besides, we consider the flux creep effects and their influence on the current re-distribution during a dwell.Comment: 13 pages, 16 figures (EPS), RevTeX4. In the revised version, corrections to perturbing effects near the weak links are introduced, one more figure is added. lin

Cell-cell adhesion regulates Merlin/NF2 interaction with the PAF complex

The PAF complex (PAFC) coordinates transcription elongation and mRNA processing and its CDC73/parafibromin subunit functions as a tumour suppressor. The NF2/Merlin tumour suppressor functions both at the cell cortex and nucleus and is a key mediator of contact inhibition but the molecular mechanisms remain unclear. In this study we have used affinity proteomics to identify novel Merlin interacting proteins and show that Merlin forms a complex with multiple proteins involved in RNA processing including the PAFC and the CHD1 chromatin remodeller. Tumour-derived inactivating mutations in both Merlin and the CDC73 PAFC subunit mutually disrupt their interaction and growth suppression by Merlin requires CDC73. Merlin interacts with the PAFC in a cell density-dependent manner and we identify a role for FAT cadherins in regulating the Merlin-PAFC interaction. Our results suggest that in addition to its function within the Hippo pathway, Merlin is part of a tumour suppressor network regulated by cell-cell adhesion which coordinates post-initiation steps of the transcription cycle of genes mediating contact inhibition

Novel Mitochondrial Substrates of Omi Indicate a New Regulatory Role in Neurodegenerative Disorders

The mitochondrial protease OMI (also known as HtrA2) has been implicated in Parkinson's Disease (PD) and deletion or protease domain point mutations have shown profound neuropathologies in mice. A beneficial role by OMI, in preserving cell viability, is assumed to occur via the avoidance of dysfunctional protein turnover. However relatively few substrates for mitochondrial Omi are known. Here we report our identification of three novel mitochondrial substrates that impact metabolism and ATP production. Using a dual proteomic approach we have identified three interactors based upon ability to bind to OMI, and/or to persist in the proteome after OMI activity has been selectively inhibited. One candidate, the chaperone HSPA8, was common to each independent study. Two others (PDHB subunit and IDH3A subunit) did not appear to bind to OMI, however persisted in the mito-proteome when OMI was inhibited. Pyruvate dehydrogenase (PDH) and isocitrate dehydrogenase (IDH) are two key Kreb's cycle enzymes that catalyse oxidative decarboxylation control points in mitochondrial respiration. We verified both PDHB and IDH3A co-immunoprecipitate with HSPA8 and after elution, were degraded by recombinant HtrA2 in vitro. Additionally our gene expression studies, using rotenone (an inhibitor of Complex I) showed Omi expression was silenced when pdhb and idh3a were increased when a sub-lethal dose was applied. However higher dose treatment caused increased Omi expression and decreased levels of pdhb and idh3a transcripts. This implicates mitochondrial OMI in a novel mechanism relating to metabolism

HtrA2/Omi Terminates Cytomegalovirus Infection and Is Controlled by the Viral Mitochondrial Inhibitor of Apoptosis (vMIA)

Viruses encode suppressors of cell death to block intrinsic and extrinsic host-initiated death pathways that reduce viral yield as well as control the termination of infection. Cytomegalovirus (CMV) infection terminates by a caspase-independent cell fragmentation process after an extended period of continuous virus production. The viral mitochondria-localized inhibitor of apoptosis (vMIA; a product of the UL37x1 gene) controls this fragmentation process. UL37x1 mutant virus-infected cells fragment three to four days earlier than cells infected with wt virus. Here, we demonstrate that infected cell death is dependent on serine proteases. We identify mitochondrial serine protease HtrA2/Omi as the initiator of this caspase-independent death pathway. Infected fibroblasts develop susceptibility to death as levels of mitochondria-resident HtrA2/Omi protease increase. Cell death is suppressed by the serine protease inhibitor TLCK as well as by the HtrA2-specific inhibitor UCF-101. Experimental overexpression of HtrA2/Omi, but not a catalytic site mutant of the enzyme, sensitizes infected cells to death that can be blocked by vMIA or protease inhibitors. Uninfected cells are completely resistant to HtrA2/Omi induced death. Thus, in addition to suppression of apoptosis and autophagy, vMIA naturally controls a novel serine protease-dependent CMV-infected cell-specific programmed cell death (cmvPCD) pathway that terminates the CMV replication cycle

Clustering of negative affectivity and social inhibition in the community: Prevalence of type D personality as a cardiovascular risk marker.

OBJECTIVE: To explore the prevalence of Type D personality-the combination of negative affectivity and social inhibition-in the general population and its relationship to other cardiovascular risk factors, including psychopathological symptoms. Type D personality has been identified as a prognostic risk factor for various cardiovascular disease conditions. METHODS: In a representative sample of 2698 individuals (aged 35-74 years), psychological, lifestyle, and somatic risk factors were investigated with laboratory testing, self-report measures, and a clinical interview. Type D was assessed with the German Type D Scale-14. RESULTS: The prevalence of Type D was 23.4% (95% confidence interval [CI], 21.2-25.6) in men and 26.9% (95% CI, 23.7-30.1) in women and, thus, in the range of classical risk factors (e.g., hypercholesterolemia). In age-adjusted analysis, Type D was associated with psychopathological symptoms, including depression and somatic symptom burden. With the exception of physical inactivity in both sexes, hypertension in women and hypercholesterolemia in men, Type D was not associated with classical cardiovascular risk factors. Multivariate analysis revealed depression, exhaustion, anxiety, and low self-rated health as associated with Type D in both sexes (odds ratios, 1.97-3.21 in men, 1.52-2.44 in women). CONCLUSIONS: A Type D personality disposition can be found in about a quarter of the general population, which is comparable to the prevalence of classical cardiovascular risk factors. In both sexes, an independent association to Type D appeared mainly in psychopathological symptoms. Type D constitutes a relevant and independent risk marker in the community and should receive attention in clinical practice

Korrelation zwischen der Realstruktur und den physikalischen Eigenschaften von HTSL-Einkristallen und -Schichten Abschlussbericht

1. Mittels Phasendiagrammuntersuchungen sowie Kristallisationsexperimenten bei unterschiedlichem Sauerstoffpartialdruck wurde das primaere Kristallisationsfeld des YBCO bestimmt und die isotherme Zuechtung ermoeglicht. 2. Es wurden aus Hochtemperaturloesungen YBCO-Einkristalle hoher kristallographischer Perfektion hergestellt und Aussagen zum Verstaendnis der Flussverankerung gewonnen. 3. Mittels Fluessigphasenepitaxie (LPE) von YBCO auf NdGaO_3- und LaGaO_3-Substraten sind c- und (international erstmalig) a,b-Schichten mit Einkritallqualitaet (Tc#approx#90 K, #DELTA#T#<=#0.5 K) hergestellt und strukturell wie auch magnetisch charakterisiert worden. 4. Erste schmelztexturierte YBCO-''Einkristalle'' haben intrakristalline Stromdichten > 10"4 A/cm"2 bei Induktionen bis zu 2 T und Rreversibilitaetsinduktionen bis 6 T bei 77 K. Zusammen mit Kooperationspartnern wurden Vorstellungen zum Wachstumsmechanismus entwickelt und publiziert. 5. Insbesondere mit torsionsmagnetometrischen Untersuchungen ist es gelungen, intrinsische von realstrukturbedingten Effekten abzutrennen und damit die Moeglichkeiten einer gezielten Beeinflussung der Pinningeigenschaften abzugrenzen. 6. Die im wesentlichen eigenstaendigen Beitraege zur Theorie magnetischer Messverfahren fuehrten zur Separation von geometrischen und apparativen Einfluessen und damit zu einer klaren Interpretation magnetischer Messungen, insbesondere der Torsionsmagnetometric und der ac-Suszeptometrie. (orig./MM)1. In order to carry out isothermic crystal growth experiments of YBCO the 123 primary crystallization field was determined by means of phase diagram investigations and crystal growth experiments at different oxygen partial pressure. 2. YBCO single crystals of high crystallographic perfection were grown and conclusions on the flux pinning mechanism were drawn. 3. By means of Liquid Phase Epitaxy (LPE) single crystalline (Tc#approx#90 K; #DELTA#T#<=#0.5 K) c-and a,b- YBCO fils have been prepared on NdGaO_3 and LaGaO_3 substrates. The films were characterized structurally and magnetically. 4. Our fist melt textured YBCO ''single crystals'' possess intracrystalline critical current densities >10"4 A/cm"2 at B#<=#2T. The irreversibility inductions are #<=#6 T at 77 K. A simple demonstrator was constructed together with the IFW Dresden and a growth model was developed. 5. Using above all torque magnetometer measurements we separated intrinsic pinning effects from effects which are reasoned by the microstructure. 6. Own contributions to the theory of magnetic measuring methods resulted in the separation of geometric influences and influences of experimental arrangement, what leads to a clear interpretation of magnetic measurements carried out mainly with torque magnetometry and ac-susceptometry. (orig./MM)Available from TIB Hannover: D.Dt.F.QN1(7,27) / FIZ - Fachinformationszzentrum Karlsruhe / TIB - Technische InformationsbibliothekSIGLEBundesministerium fuer Forschung und Technologie (BMFT), Bonn (Germany)DEGerman