Identification of DNA damage-induced autophagic cargo by mass-spectrometry based proteomics

Autophagy is a catabolic recycling pathway initiated under periods of starvation with energy-saving and cytoprotective effects. Previously considered a non-targeted process that degrades bulk cytoplasmic material, the discovery of autophagy receptors revealed that autophagy also leads to the targeted turnover of different cargo. Upon its perception, DNA damage triggers tailor-made signaling and repair pathways, collectively designated as the DNA damage response (DDR) that can also result in apoptosis or cellular senescence. Kinases that orchestrate the DDR are ATM, ATR and DNA-PKcs. Autophagy succeeds DNA damage, but the signaling pathways required to transmit the information for autophagy activation from the nucleus to the cytoplasm as well as its purpose remain poorly understood. This study focused on two aspects of DNA damage-induced autophagy. First, I investigated whether the DDR kinases play a role in the activation of autophagy after DNA double strand breaks. Second, I employed mass spectrometry (MS)-based proteomics to probe the cargo of DNA damage-induced autophagy to investigate its relevance in the DDR. To assess the role of the DDR kinases in DNA damage-induced autophagy, the conversion of the autophagy marker protein LC3B into its lipidated form and the degradation of the receptor p62 were monitored in DDR kinase knockdown cells. As a complementary approach, the tandem-LC3B assay was engaged to measure DNA damage-induced autophagy in the presence of DDR kinase inhibitors. Using different read-out systems, we came to indecisive results indicating no, mild or strong involvement of the DDR kinases in DNA damage-induced autophagy. The off-target effects of commonly used DDR kinase inhibitors that might also cause the inhibition of the phosphatidylinositol-3-kinase VPS34 could explain observed inconsistency between the results obtained in knockdown and chemical inhibitor-treated cells. I established a MS-based proximity proteomics approach to identify systematically DNA damage-induced autophagic cargo. Applying this technique, many known and previously undescribed putative cargo proteins were identified. Computational analyses permitted to associate the identified cargo with focal adhesions proteins, ribosomal proteins, the subunits of the TRiQ/CCT chaperonin complex as well as with nuclear-localized proteins. The proteinase K protection assay enabled to validate most of the selected proteins as autophagic cargo. Ultimately, the cargo that was identified indicates that micronuclei might be degraded by autophagy after genotoxic stress. Further, identification of focal adhesions proteins and validation of the metastasis suppressor NME1 could indicate for a metastasis-promoting role of autophagy after treatment with chemotherapeutic agents. This study presents the first approach to investigate DNA damage-induced autophagic cargo using proximity proteomics and reveals a role for autophagy in the degradation of nuclear proteins in human cells.V, 107 Seiten, Illustrationen, Diagramm

High-pressure synthesis of superconducting Nb{1-x}B2 (x = 0-0.48) with the maximum Tc = 9.2 K

Superconductivity with Tc above 9 K was found in metal-deficient NbB2 prepared under 5 GPa, while no clear superconductivity was observed down to 3 K in stoichiometric NbB2. The superconductivity was observed above x = 0.04 in Nb1-xB2. and the lattice parameters also changed abruptly at x = 0.04. As x increased, the transition temperature Tc slightly rose and fell with the maximum value of 9.2 K at x = 0.24 for the samples sintered at 5 GPa and 1200 C. The Tc-value changed in the range from 7 K to 9 K, depending on the sintering pressure. A series of Ta1-xB2 (0 =< x =< 0.24) was also synthesized under high pressure to examine a special effect of high-pressure synthesis.Comment: 24 pages including 2 tables and 7 figures, accepted for publication in Physica

Hole Superconductivity in MgB2Mg B_2: a high TcT_c cuprate without Cu

The theory of hole superconductivity explains high temperature superconductivity in cuprates as driven by pairing of hole carriers in oxygen $p\pi$ orbitals in the highly negatively charged $Cu-O$ planes. The pairing mechanism is hole undressing and is Coulomb-interaction driven. We propose that the planes of $B$ atoms in $Mg B_2$ are akin to the $Cu-O$ planes without $Cu$, and that the recently observed high temperature superconductivity in $Mg B_2$ arises similarly from undressing of hole carriers in the planar boron $p_{x,y}$ orbitals in the negatively charged $B^-$ planes. Doping $Mg B_2$ with electrons and with holes should mirror the behavior of underdoped and overdoped high $T_c$ cuprates respectively. We discuss possible ways to achieve higher transition temperatures in boron compounds based on this theory.Comment: A section on isotope effect has been added, as well as other minor change

Electron and field interactions with thin films and surfaces Final report

Electron and field interactions with thin films and surface

Recent Progress in the Computational Many-Body Theory of Metal Surfaces

In this article we describe recent progress in the computational many-body theory of metal surfaces, and focus on current techniques beyond the local-density approximation of density-functional theory. We overview various applications to ground and excited states. We discuss the exchange-correlation hole, the surface energy, and the work function of jellium surfaces, as obtained within the random-phase approximation, a time-dependent density-functional approach, and quantum Monte Carlo methods. We also present a survey of recent quasiparticle calculations of unoccupied states at both jellium and real surfaces.Comment: 17 pages, 1 figure, to appear in Comput. Phys. Commu

Las corrientes prerrománticas en el curso del siglo XVII.

Sin resume

Heterogeneity and Disorder: Contributions of Rolf Landauer

Rolf Landauer made important contributions to many branches of science. Within the broad area of transport in disordered media, he wrote seminal papers on electrical conduction in macroscopically inhomogeneous materials, as well as fundamental analyses of electron transport in quantum mechanical systems with disorder on the atomic scale. We review here some of these contributions. We also briefly describe some main events in his personal and scientific life.Comment: 10 pages, 3 figures; presented on the occasion when Rolf Landauer was awarded, posthumously, the inaugural ETOPIM Medal at the ETOPIM 8 Conference, which took place during 7--12 June, 2009 in Rethymnon, Cret

Genetic requirements for repair of lesions caused by single genomic ribonucleotides in S phase

Single ribonucleoside monophosphates (rNMPs) are transiently present in eukaryotic genomes. The RNase H2-dependent ribonucleotide excision repair (RER) pathway ensures error-free rNMP removal. In some pathological conditions, rNMP removal is impaired. If these rNMPs hydrolyze during, or prior to, S phase, toxic single-ended double-strand breaks (seDSBs) can occur upon an encounter with replication forks. How such rNMP-derived seDSB lesions are repaired is unclear. We expressed a cell cycle phase restricted allele of RNase H2 to nick at rNMPs in S phase and study their repair. Although Top1 is dispensable, the RAD52 epistasis group and Rtt101Mms1-Mms22 dependent ubiquitylation of histone H3 become essential for rNMP-derived lesion tolerance. Consistently, loss of Rtt101Mms1-Mms22 combined with RNase H2 dysfunction leads to compromised cellular fitness. We refer to this repair pathway as nick lesion repair (NLR). The NLR genetic network may have important implications in the context of human pathologies