Exosomes released from breast cancer carcinomas stimulate cell movement

For metastasis to occur cells must communicate with to their local environment to initiate growth and invasion. Exosomes have emerged as an important mediator of cell-to-cell signalling through the transfer of molecules such as mRNAs, microRNAs, and proteins between cells. Exosomes have been proposed to act as regulators of cancer progression. Here, we study the effect of exosomes on cell migration, an important step in metastasis. We performed cell migration assays, endocytosis assays, and exosome proteomic profiling on exosomes released from three breast cancer cell lines that model progressive stages of metastasis. Results from these experiments suggest: (1) exosomes promote cell migration and (2) the signal is stronger from exosomes isolated from cells with higher metastatic potentials; (3) exosomes are endocytosed at the same rate regardless of the cell type; (4) exosomes released from cells show differential enrichment of proteins with unique protein signatures of both identity and abundance. We conclude that breast cancer cells of increasing metastatic potential secrete exosomes with distinct protein signatures that proportionally increase cell movement and suggest that released exosomes could play an active role in metastasis

SIRT3‐dependent deacetylation exacerbates acetaminophen hepatotoxicity

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/102226/1/embr2011121.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/102226/2/embr2011121-sup-0001.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/102226/3/embr2011121.reviewer_comments.pd

Biotin ligase tagging identifies proteins proximal to E-cadherin, including lipoma preferred partner, a regulator of epithelial cell–cell and cell–substrate adhesion

Known proteins associated with the cell-adhesion protein E-cadherin include catenins and proteins involved in signaling, trafficking and actin organization. However, the list of identified adherens junction proteins is likely to be incomplete, limiting investigation into this essential cell structure. To expand the inventory of potentially relevant proteins, we expressed E-cadherin fused to biotin ligase in MDCK epithelial cells, and identified by mass spectrometry neighboring proteins that were biotinylated. The most abundant of the 303 proteins identified were catenins and nearly 40 others that had been previously reported to influence cadherin function. Many others could be rationalized as novel candidates for regulating the adherens junction, cytoskeleton, trafficking or signaling. We further characterized lipoma preferred partner (LPP), which is present at both cell contacts and focal adhesions. Knockdown of LPP demonstrated its requirement for E-cadherin-dependent adhesion and suggested that it plays a role in coordination of the cell–cell and cell–substrate cytoskeletal interactions. The analysis of LPP function demonstrates proof of principle that the proteomic analysis of E-cadherin proximal proteins expands the inventory of components and tools for understanding the function of E-cadherin

Proteomics approaches to fibrotic disorders

This review provides an introduction to mass spectrometry based proteomics and discusses several proteomics approaches that are relevant in understanding the pathophysiology of fibrotic disorders and the approaches that are frequently used in biomarker discovery

A Mighty Small Heart: The Cardiac Proteome of Adult Drosophila melanogaster

Drosophila melanogaster is emerging as a powerful model system for the study of cardiac disease. Establishing peptide and protein maps of the Drosophila heart is central to implementation of protein network studies that will allow us to assess the hallmarks of Drosophila heart pathogenesis and gauge the degree of conservation with human disease mechanisms on a systems level. Using a gel-LC-MS/MS approach, we identified 1228 protein clusters from 145 dissected adult fly hearts. Contractile, cytostructural and mitochondrial proteins were most abundant consistent with electron micrographs of the Drosophila cardiac tube. Functional/Ontological enrichment analysis further showed that proteins involved in glycolysis, Ca2+-binding, redox, and G-protein signaling, among other processes, are also over-represented. Comparison with a mouse heart proteome revealed conservation at the level of molecular function, biological processes and cellular components. The subsisting peptidome encompassed 5169 distinct heart-associated peptides, of which 1293 (25%) had not been identified in a recent Drosophila peptide compendium. PeptideClassifier analysis was further used to map peptides to specific gene-models. 1872 peptides provide valuable information about protein isoform groups whereas a further 3112 uniquely identify specific protein isoforms and may be used as a heart-associated peptide resource for quantitative proteomic approaches based on multiple-reaction monitoring. In summary, identification of excitation-contraction protein landmarks, orthologues of proteins associated with cardiovascular defects, and conservation of protein ontologies, provides testimony to the heart-like character of the Drosophila cardiac tube and to the utility of proteomics as a complement to the power of genetics in this growing model of human heart disease

Barrier-to-Autointegration Factor Proteome Reveals Chromatin-Regulatory Partners

Nuclear lamin filaments and associated proteins form a nucleoskeletal (“lamina”) network required for transcription, replication, chromatin organization and epigenetic regulation in metazoans. Lamina defects cause human disease (“laminopathies”) and are linked to aging. Barrier-to-autointegration factor (BAF) is a mobile and essential component of the nuclear lamina that binds directly to histones, lamins and LEM-domain proteins, including the inner nuclear membrane protein emerin, and has roles in chromatin structure, mitosis and gene regulation. To understand BAF's mechanisms of action, BAF associated proteins were affinity-purified from HeLa cell nuclear lysates using BAF-conjugated beads, and identified by tandem mass spectrometry or independently identified and quantified using the iTRAQ method. We recovered A- and B-type lamins and core histones, all known to bind BAF directly, plus four human transcription factors (Requiem, NonO, p15, LEDGF), disease-linked proteins (e.g., Huntingtin, Treacle) and several proteins and enzymes that regulate chromatin. Association with endogenous BAF was independently validated by co-immunoprecipitation from HeLa cells for seven candidates including Requiem, poly(ADP-ribose) polymerase 1 (PARP1), retinoblastoma binding protein 4 (RBBP4), damage-specific DNA binding protein 1 (DDB1) and DDB2. Interestingly, endogenous BAF and emerin each associated with DDB2 and CUL4A in a UV- and time-dependent manner, suggesting BAF and emerin have dynamic roles in genome integrity and might help couple DNA damage responses to the nuclear lamina network. We conclude this proteome is a rich source of candidate partners for BAF and potentially also A- and B-type lamins, which may reveal how chromatin regulation and genome integrity are linked to nuclear structure

Global discovery of high-NaCl-induced changes of protein phosphorylation

U ovoj su radnji izvršene analize novih pristupa u prenaponskoj zaštiti srednjenaponskih elektroenergetskih mreža. Analizirani su nastanci i djelovanja prenapona, te je dan pregled postojećih metoda prenaponske zaštite u srednjenaponskim distribucijskim mrežama. Kao novi pristupi u prenaponskoj zaštiti, analizirani su dugi preskočni odvodnik prenapona s antenskim modulom, te najnovija saznanja o uzrocima i zaštitnim mjerama na polju prenaponske zaštite uslijed djelovanja induciranih napona. Predstavljen je princip djelovanja LFA-AM (Long Flashover Arrester with Antenna-module) dugog preskočnog odvodnika prenapona s antenskim modulom, te izvršena analiza laboratorijskih testova. Prenaponska zaštita od pojave previsokih induciranih napona je obrađena od analize metoda proračuna induciranih napona, preko utjecaja zaštitnog vodiča na vodu i susjednih objekata na učestalost i veličinu induciranih napona, do prijedloga zaštite i usporedbe s vrijednostima dobivenima u ispitivanju na terenu. Dan je poseban osvrt na stanje prenaponske zaštite u srednjenaponskom elektroenergetskom sustavu EPHZHB.In this master thesis, a new approaches to overvoltage protection of middlevoltage power networks are carried out. Overvoltage generation and its effects are analysed and also an overview of existing methods of overvoltage protection in middle-voltage power networks is presented. As new approaches to overvoltage protection, Long Flashover Arrester with Antenna-module (LFA-AM) and the newest cognitions about causes and protection measures on the area of protection against induced voltages are analysed. Operating principle and design of Long Flashover Arrester with Antenna-module(LFA-AM) are presented and appropriate laboratory tests are performed. Overvoltage protection against induced voltages is done through: analysis of calculation methods of induced voltages, analysis of ground wire and nearby objects effect on occurrence frequency and values of induced voltages, suggestions of overvoltage protection measures and comparison of calculated results to field data. A special overview of overvoltage protection of middle-voltage power networks in EPHZHB power electricity company is given