Specific Set Of Proteins Research Articles

OGA mutant aberrantly hydrolyzes O-GlcNAc modification from PDLIM7 to modulate p53 and cytoskeleton in promoting cancer cell malignancy

O-GlcNAcase (OGA) is the only human enzyme that catalyzes the hydrolysis (deglycosylation) of O-linked beta-N-acetylglucosaminylation (O-GlcNAcylation) from numerous protein substrates. OGA has broad implications in many challenging diseases including cancer. However, its role in cell malignancy remains mostly unclear. Here, we report that a cancer-derived point mutation on the OGA's noncatalytic stalk domain aberrantly modulates OGA interactome and substrate deglycosylation toward a specific set of proteins. Interestingly, our quantitative proteomic studies uncovered that the OGA stalk domain mutant preferentially deglycosylated protein substrates with +2 proline in the sequence relative to the O-GlcNAcylation site. One of the most dysregulated substrates is PDZ and LIM domain protein 7 (PDLIM7), which is associated with the tumor suppressor p53. We found that the aberrantly deglycosylated PDLIM7 suppressed p53 gene expression and accelerated p53 protein degradation by promoting the complex formation with E3 ubiquitin ligase MDM2. Moreover, deglycosylated PDLIM7 significantly up-regulated the actin-rich membrane protrusions on the cell surface, augmenting the cancer cell motility and aggressiveness. These findings revealed an important but previously unappreciated role of OGA's stalk domain in protein substrate recognition and functional modulation during malignant cell progression.

Proteomic, Metabolomic, and Fatty Acid Profiling of Small Extracellular Vesicles from Glioblastoma Stem-Like Cells and Their Role in Tumor Heterogeneity.

Glioblastoma is a deadly brain tumor for which there is no cure. The presence of glioblastoma stem-like cells (GSCs) contributes to the heterogeneous nature of the disease and makes developing effective therapies challenging. Glioblastoma cells have been shown to influence their environment by releasing biological nanostructures known as extracellular vesicles (EVs). Here, we investigated the role of GSC-derived nanosized EVs (<200 nm) in glioblastoma heterogeneity, plasticity, and aggressiveness, with a particular focus on their protein, metabolite, and fatty acid content. We showed that conditioned medium and small extracellular vesicles (sEVs) derived from cells of one glioblastoma subtype induced transcriptomic and proteomic changes in cells of another subtype. We found that GSC-derived sEVs are enriched in proteins playing a role in the transmembrane transport of amino acids, carboxylic acids, and organic acids, growth factor binding, and metabolites associated with amino acid, carboxylic acid, and sugar metabolism. This suggests a dual role of GSC-derived sEVs in supplying neighboring GSCs with valuable metabolites and proteins responsible for their transport. Moreover, GSC-derived sEVs were enriched in saturated fatty acids, while their respective cells were high in unsaturated fatty acids, supporting that the loading of biological cargos into sEVs is a highly regulated process and that GSC-derived sEVs could be sources of saturated fatty acids for the maintenance of glioblastoma cell metabolism. Interestingly, sEVs isolated from GSCs of the proneural and mesenchymal subtypes are enriched in specific sets of proteins, metabolites, and fatty acids, suggesting a molecular collaboration between transcriptionally different glioblastoma cells. In summary, this study revealed the complexity of GSC-derived sEVs and unveiled their potential contribution to tumor heterogeneity and critical cellular processes commonly deregulated in glioblastoma.

Kinesin-1 patterns Par-1 and Rho signaling at the cortex of syncytial embryos of Drosophila.

The cell cortex of syncytial Drosophila embryos is patterned into cap and intercap regions by centrosomes, specific sets of proteins that are restricted to their respective regions by unknown mechanisms. Here, we found that Kinesin-1 is required for the restriction of plus- and minus-ends of centrosomal and non-centrosomal microtubules to the cap region, marked by EB1 and Patronin/Shot, respectively. Kinesin-1 also directly or indirectly restricts proteins and Rho signaling to the intercap, including the RhoGEF Pebble, Dia, Myosin II, Capping protein-α, and the polarity protein Par-1. Furthermore, we found that Par-1 is required for cap restriction of Patronin/Shot, and vice versa Patronin, for Par-1 enrichment at the intercap. In summary, our data support a model that Kinesin-1 would mediate the restriction of centrosomal and non-centrosomal microtubules to a region close to the centrosomes and exclude Rho signaling and Par-1. In addition, mutual antagonistic interactions would refine and maintain the boundary between cap and intercap and thus generate a distinct cortical pattern.

A myeloid leukemia factor homolog is involved in tolerance to stresses and stress-induced protein metabolism in Giardia lamblia

BackgroundThe eukaryotic membrane vesicles contain specific sets of proteins that determine vesicle function and shuttle with specific destination. Giardia lamblia contains unknown cytosolic vesicles that are related to the identification of a homolog of human myeloid leukemia factor (MLF) named MLF vesicles (MLFVs). Previous studies suggest that MLF also colocalized with two autophagy machineries, FYVE and ATG8-like protein, and that MLFVs are stress-induced compartments for substrates of the proteasome or autophagy in response to rapamycin, MG132, and chloroquine treatment. A mutant protein of cyclin-dependent kinase 2, CDK2m3, was used to understand whether the aberrant proteins are targeted to degradative compratments. Interestingly, MLF was upregulated by CDK2m3 and they both colocalized within the same vesicles. Autophagy is a self-digestion process that is activated to remove damaged proteins for preventing cell death in response to various stresses. Because of the absence of some autophagy machineries, the mechanism of autophagy is unclear in G. lamblia.ResultsIn this study, we tested the six autophagosome and stress inducers in mammalian cells, including MG132, rapamycin, chloroquine, nocodazole, DTT, and G418, and found that their treatment increased reactive oxygen species production and vesicle number and level of MLF, FYVE, and ATG8-like protein in G. lamblia. Five stress inducers also increased the CDK2m3 protein levels and vesicles. Using stress inducers and knockdown system for MLF, we identified that stress induction of CDK2m3 was positively regulated by MLF. An autophagosome-reducing agent, 3-methyl adenine, can reduce MLF and CDK2m3 vesicles and proteins. In addition, knockdown of MLF with CRISPR/Cas9 system reduced cell survival upon treatment with stress inducers. Our newly developed complementation system for CRISPR/Cas9 indicated that complementation of MLF restored cell survival in response to stress inducers. Furthermore, human MLF2, like Giardia MLF, can increase cyst wall protein expression and cyst formation in G. lamblia, and it can colocalize with MLFVs and interact with MLF.ConclusionsOur results suggest that MLF family proteins are functionally conserved in evolution. Our results also suggest an important role of MLF in survival in stress conditions and that MLFVs share similar stress-induced characteristics with autophagy compartments.

Quantitative proteomic analysis of skeletal muscles from wild-type and transgenic mice carrying recessive Ryr1 mutations linked to congenital myopathies.

Skeletal muscles are a highly structured tissue responsible for movement and metabolic regulation, which can be broadly subdivided into fast and slow twitch muscles with each type expressing common as well as specific sets of proteins. Congenital myopathies are a group of muscle diseases leading to a weak muscle phenotype caused by mutations in a number of genes including RYR1. Patients carrying recessive RYR1 mutations usually present from birth and are generally more severely affected, showing preferential involvement of fast twitch muscles as well as extraocular and facial muscles. In order to gain more insight into the pathophysiology of recessive RYR1-congential myopathies, we performed relative and absolute quantitative proteomic analysis of skeletal muscles from wild-type and transgenic mice carrying p.Q1970fsX16 and p.A4329D RyR1 mutations which were identified in a child with a severe congenital myopathy. Our in-depth proteomic analysis shows that recessive RYR1 mutations not only decrease the content of RyR1 protein in muscle, but change the expression of 1130, 753, and 967 proteins EDL, soleus and extraocular muscles, respectively. Specifically, recessive RYR1 mutations affect the expression level of proteins involved in calcium signaling, extracellular matrix, metabolism and ER protein quality control. This study also reveals the stoichiometry of major proteins involved in excitation contraction coupling and identifies novel potential pharmacological targets to treat RyR1-related congenital myopathies.

Normal oil body formation in Marchantia polymorpha requires functional coat protein complex I proteins.

Eukaryotic cells possess endomembrane organelles equipped with specific sets of proteins, lipids, and polysaccharides that are fundamental for realizing each organelle’s specific function and shape. A tightly regulated membrane trafficking system mediates the transportation and localization of these substances. Generally, the secretory/exocytic pathway is responsible for transporting cargo to the plasma membrane and/or the extracellular space. However, in the case of oil body cells in the liverwort Marchantia polymorpha, the oil body, a liverwort-unique organelle, is thought to be formed by secretory vesicle fusion through redirection of the secretory pathway inside the cell. Although their formation mechanism remains largely unclear, oil bodies exhibit a complex and bumpy surface structure. In this study, we isolated a mutant with spherical oil bodies through visual screening of mutants with abnormally shaped oil bodies. This mutant harbored a mutation in a coat protein complex I (COPI) subunit MpSEC28, and a similar effect on oil body morphology was also detected in knockdown mutants of other COPI subunits. Fluorescently tagged MpSEC28 was localized to the periphery of the Golgi apparatus together with other subunits, suggesting that it is involved in retrograde transport from and/or in the Golgi apparatus as a component of the COPI coat. The Mpsec28 mutants also exhibited weakened stiffness of the thalli, suggesting impaired cell–cell adhesion and cell wall integrity. These findings suggest that the mechanism of cell wall biosynthesis is also involved in shaping the oil body in M. polymorpha, supporting the redirection of the secretory pathway inward the cell during oil body formation.

Decoding the Oncogenic Signals from the Long Non-Coding RNAs

Cancer is one of the leading causes of death worldwide. Multifactorial etiology of cancer and tumor heterogeneity are the two most acute challenges in existing diagnostic and therapeutic strategies for cancer. An effective precision cancer medicine strategy to overcome these challenges requires a clear understanding of the transcriptomic landscape of cancer cells. Recent innovative breakthroughs in high-throughput sequencing technologies have identified the oncogenic or tumor-suppressor role of several long non-coding RNAs (lncRNAs). LncRNAs have been characterized as regulating various signaling cascades which are involved in the pathobiology of cancer. They modulate cancer cell survival, proliferation, metabolism, invasive metastasis, stemness, and therapy-resistance through their interactions with specific sets of proteins, miRNAs and other non-coding RNAs, mRNAs, or DNAs in cells. By virtue of their ability to regulate multiple sets of genes and their cognate signaling pathways, lncRNAs are emerging as potential candidates for diagnostic, prognostic, and therapeutic targets. This review is focused on providing insight into the mechanisms by which different lncRNAs play a critical role in cancer growth, and their potential role in cancer diagnosis, prognosis, and therapy.

Deciphering ploidal levels of Lippia alba by using proteomics

Lippia alba (Mill.) N.E. Brown (Verbenaceae), popularly known as “lemon balm” or “bushy matgrass”, is widely used in folk medicine due to its anti-inflammatory, antispasmodic, analgesic, and digestive properties. It was described as an autopolyploid complex with five cytotypes (2n = 30, 38, 45, 60 and 90). To enhance our understanding of the biological variation of the species, we investigated, comparatively, the proteomic profile of all ploidal levels (diploid, aneuploid, triploid, tetraploid, and hexaploid). Leaf proteins were extracted with subsequent separation by two-dimensional electrophoresis, spot analysis, and protein identification by mass spectrometry. By comparing the proteomic profile of diploid accession to the profile of the other ploidal levels we identified differential expression between the analysed spots. We identified 34 proteins with differential expression between the ploidal levels in comparison with the diploid. The identified proteins seem to play relevant roles in the primary metabolism of L. alba suggesting that a specific set of proteins was selected during the polyploidization process, being the triploid the most different one. Given that protein composition can substantially affect the desired therapeutic effect, we posit that further combination of proteomic and metabolomic studies may help to unravel genetic variations and phenotypic profiles in L. alba.

Lipid droplets form a network interconnected by the endoplasmic reticulum through which their proteins equilibrate.

Lipid droplets (LDs) are globular intracellular structures dedicated to the storage of neutral lipids. They are closely associated with the endoplasmic reticulum (ER) and are delineated by a monolayer of phospholipids that is continuous with the cytoplasmic leaflet of the ER membrane. LDs contain a specific set of proteins, but how these proteins are targeted to the LD surface is not fully understood. Here, we devised a yeast mating-based microscopic readout to monitor the transfer of LD proteins upon zygote formation. The results of this analysis indicate that ER fusion between mating partners is required for transfer of LD proteins and that this transfer is continuous, bidirectional and affects most LDs simultaneously. These observations suggest that LDs do not fuse upon mating of yeast cells, but that they form a network that is interconnected through the ER membrane. Consistent with this, ER-localized LD proteins rapidly move onto LDs of a mating partner and this protein transfer is affected by seipin, a protein important for proper LD biogenesis and the functional connection of LDs with the ER membrane.

Specific Seminal Plasma Fractions Are Responsible for the Modulation of Sperm-PMN Binding in the Donkey.

Simple SummaryPrevious research demonstrated that seminal plasma, the fluid that together with sperm composes the semen, modulates the interaction between sperm and polymorphonuclear neutrophils (PMN) within the donkey uterus. This study sought to evaluate whether a specific set of proteins or the entire seminal plasma is involved in that modulation. Seminal plasma was fractioned into six portions: <3, 3–10, 10–30, 30–50, 50–100, and >100 kDa. Sperm incubated with two specific fractions, containing 30–50 kDa and 50–100 kDa proteins, exhibited the highest motility and viability after co-incubation with PMN for 4 h, and the highest sperm–PMN interaction at the beginning of the experiment. These results suggest that proteins of 30–50 and 50–100 kDa, rather than the entire seminal plasma, appear to be involved in the inflammatory reaction within the donkey endometrium.While artificial insemination (AI) with frozen-thawed sperm results in low fertility rates in donkeys, the addition of seminal plasma, removed during cryopreservation, partially counteracts that reduction. Related to this, an apparent inflammatory reaction in jennies is induced following AI with frozen-thawed sperm, as a high amount of polymorphonuclear neutrophils (PMN) are observed within the donkey uterus six hours after AI. While PMN appear to select the sperm that ultimately reach the oviduct, two mechanisms, phagocytosis and NETosis, have been purported to be involved in that clearance. Remarkably, sperm interacts with PMN, but the presence of seminal plasma reduces that binding. As seminal plasma is a complex fluid made up of different molecules, including proteins, this study aimed to evaluate how different seminal plasma fractions, separated by molecular weight (<3, 3–10, 10–30, 30–50, 50–100, and >100 kDa), affect sperm–PMN binding. Sperm motility, viability, and sperm–PMN binding were evaluated after 0 h, 1 h, 2 h, 3 h, and 4 h of co-incubation at 38 °C. Two seminal plasma fractions, including 30–50 kDa or 50–100 kDa proteins, showed the highest sperm motility and viability. As viability of sperm not bound to PMN after 3 h of incubation was the highest in the presence of 30–50 and 50–100 kDa proteins, we suggest that both fractions are involved in the control of the jenny’s post-breeding inflammatory response. In conclusion, this study has shown for the first time that specific fractions rather than the entire seminal plasma modulate sperm–PMN binding within the donkey uterus. As several proteins suggested to be involved in the control of post-AI endometritis have a molecular weight between 30 and 100 kDa, further studies aimed at determining the identity of these molecules and evaluating their potential effect in vivo are much warranted.

Chromosome Mapping of U2 snDNA in Species of Leptodactylus (Anura, Leptodactylidae)

Small nuclear RNA (snRNA) is a class of molecules involved in the processing of pre-mRNA and in regulatory cell processes. snRNAs are always associated with a set of specific proteins. The complexes are referred to as small nuclear ribonucleoproteins, and spliceosome U RNAs are their most common snRNA components. The repetitive sequences of U snDNAs have been cytogenetically mapped in several species of Arthropoda, fishes, and mammals; however, their distribution remains unknown in amphibians. Here, we show results of FISH mapping of U2 snDNA repetitive sequences in species of the amphibian genus Leptodactylus to reveal the distribution patterns of this sequence in their karyotypes. The probe hybridized in the metacentric chromosome pair 6 in Leptodactylus fuscus, L. gracilis, L. latrans, L. chaquensis, L. petersii, L. podicipinus, and L. brevipes. A different pattern was observed in L. labyrinthicus with hybridization signals in 4 chromosome pairs. The same localization of U2 gene sequences in most of the species analyzed suggests a relatively conserved pattern and a similarity of the chromosome 6 among these species of Leptodactylus.

Direct unfolding of RuvA-HJ complex at the single-molecule level

The repair of double-stranded DNA breaks via homologous recombination involves a four-way cross-strand intermediate known as Holliday junction (HJ), which is recognized, processed, and resolved by a specific set of proteins. RuvA, a prokaryotic HJ-binding protein, is known to stabilize the square-planar conformation of the HJ, which is otherwise a short-lived intermediate. Despite much progress being made regarding the molecular mechanism of RuvA-HJ interactions, the mechanochemical aspect of this protein-HJ complex is yet to be investigated. Here, we employed an optical-tweezers-based, single-molecule manipulation assay to detect the formation of RuvA-HJ complex and determined its mechanical and thermodynamic properties in a manner that would be impossible with traditional ensemble techniques. We found that the binding of RuvA increases the unfolding force (Funfold) of the HJ by ∼2-fold. Compared with the ΔGunfold of the HJ alone (54 ± 13 kcal/mol), the increased free energy of the RuvA-HJ complex (101 ± 20 kcal/mol) demonstrates that the RuvA protein stabilizes HJs. Interestingly, the protein remains bound to the mechanically melted HJ, facilitating its refolding at an unusually high force when the stretched DNA molecule is relaxed. These results suggest that the RuvA protein not only stabilizes the HJs but also induces refolding of the HJs. The single-molecule platform that we employed here for studying the RuvA-HJ interaction is broadly applicable to study other HJ-binding proteins involved in the critical DNA repair process.

Local cyclic adenosine monophosphate signalling cascades-Roles and targets in chronic kidney disease.

The molecular mechanisms underlying chronic kidney disease (CKD) are poorly understood and treatment options are limited, a situation underpinning the need for elucidating the causative molecular mechanisms and for identifying innovative treatment options. It is emerging that cyclic 3',5'-adenosine monophosphate (cAMP) signalling occurs in defined cellular compartments within nanometre dimensions in processes whose dysregulation is associated with CKD. cAMP compartmentalization is tightly controlled by a specific set of proteins, including A-kinase anchoring proteins (AKAPs) and phosphodiesterases (PDEs). AKAPs such as AKAP18, AKAP220, AKAP-Lbc and STUB1, and PDE4 coordinate arginine-vasopressin (AVP)-induced water reabsorption by collecting duct principal cells. However, hyperactivation of the AVP system is associated with kidney damage and CKD. Podocyte injury involves aberrant AKAP signalling. cAMP signalling in immune cells can be local and slow the progression of inflammatory processes typical for CKD. A major risk factor of CKD is hypertension. cAMP directs the release of the blood pressure regulator, renin, from juxtaglomerular cells, and plays a role in Na+ reabsorption through ENaC, NKCC2 and NCC in the kidney. Mutations in the cAMP hydrolysing PDE3A that cause lowering of cAMP lead to hypertension. Another major risk factor of CKD is diabetes mellitus. AKAP18 and AKAP150 and several PDEs are involved in insulin release. Despite the increasing amount of data, an understanding of functions of compartmentalized cAMP signalling with relevance for CKD is fragmentary. Uncovering functions will improve the understanding of physiological processes and identification of disease-relevant aberrations may guide towards new therapeutic concepts for the treatment of CKD.

Imperative role of sugar signaling and transport during drought stress responses in plants.

Cellular sugar status is essentially maintained during normal growth conditions but is impacted negatively during various environmental perturbations. Drought presents one such unfavorable environmental cue that hampers the photosynthetic fixation of carbon into sugars and affects their transport by lowering the cellular osmotic potential. The transport of cellular sugar is facilitated by a specific set of proteins known as sugar transporters. These transporter proteins are the key determinant of influx/ efflux of various sugars and their metabolite intermediates that support the plant growth and developmental process. Abiotic stress and especially drought stress-mediated injury results in reprogramming of sugar distribution across the cellular and subcellular compartments. Here, we have reviewed the imperative role of sugar accumulation, signaling, and transport under typical and atypical stressful environments. We have discussed the physiological effects of drought on sugar accumulation and transport through different transporter proteins involved in monosaccharide and disaccharide sugar transport. Further, we have illustrated sugar-mediated signaling and regulation of sugar transporter proteins along with the overall crosstalk of this signaling with the phytohormone module of abiotic stress response under osmotic stress. Overall, the present review highlights the critical role of sugar transport, distribution and signaling in plants under drought stress conditions.

96-OR: Integrative Omics Analysis Revealed That the Metabolic Changes in Placental Cells from Gestational Diabetes Mellitus Are Reflected in Their Secreted Exosomes

Exosomes are membrane-bound extracellular vesicles secreted from a range of cells including human placental cells and contain specific cargo such as proteins, and miRNAs. The aim of this study was to determine the miRNA and protein profile in primary human trophoblast (PTH) and their secreted exosomes obtained from placentae from women with normal glucose tolerant (NGT) and gestational diabetes mellitus (GDM). Using quantitative proteomic analysis, 34 proteins were significantly different in PTH from GDM compared with NGT. Similarly, a specific set of proteins were significantly different in PTH exosomes from GDM compared with NGT. An Illumina TrueSeq Small RNA kit was used to construct a small RNA library from PTH and exosomes. We identified a set of miRNAs (miR-181, miR-151, miR-10a, miR-744, miR-1468 and miR-4507) which their expression varied in a consistent pattern in PTH and their secreted exosomes in GDM compared with NGT. Interestingly, a specific set of proteins and miRNAs were selectively enriched with exosomes and compared with their PTH of origin in NGT and GDM indicating a specific packaging of proteins and miRNAs into exosomes. We used Ingenuity Pathway Analysis (IPA) analysis to identify miRNA-gene interactions using the miRNA and proteomic data and created a network link with the down- and up-regulated pairs of miRNA-proteins. IPA core analysis showed that the top canonical pathway associated with these miRNAs were PI3/AKT signalling and glucose metabolism/insulin resistance, respectively. We proposed that exosomes are “fingerprints” of the releasing cells and their metabolic status. Exosomes released from placenta can modify the phenotype of target cells, inducing metabolic changes in GDM to contribute to changes in insulin sensitivity during pregnancy. Disclosure S. Nair: None. N. Jayabalan: None. D. Guanzon: None. A. Lai: None. D. McIntyre: Other Relationship; Self; Novo Nordisk A/S. M. Lappas: None. C. Salomon: None. Funding Diabetes Australia; Fondo Nacional de Desarrollo Científico y Tecnológico (1170809)

Tunneling nanotube-mediated intercellular vesicle and protein transfer in the stroma-provided imatinib resistance in chronic myeloid leukemia cells

Intercellular communication within the bone marrow niche significantly promotes leukemogenesis and provides protection of leukemic cells from therapy. Secreted factors, intercellular transfer of mitochondria and the receptor–ligand interactions have been shown as mediators of this protection. Here we report that tunneling nanotubes (TNTs)—long, thin membranous structures, which have been identified as a novel mode of intercellular cross-talk—are formed in the presence of stroma and mediate transfer of cellular vesicles from stroma to leukemic cells. Importantly, transmission of vesicles via TNTs from stromal cells increases resistance of leukemic cells to the tyrosine kinase inhibitor, imatinib. Using correlative light-electron microscopy and electron tomography we show that stromal TNTs contain vesicles, provide membrane continuity with the cell bodies and can be open-ended. Moreover, trans-SILAC studies to reveal the non-autonomous proteome showed that specific sets of proteins are transferred together with cellular vesicles from stromal to leukemic cells, with a potential role in survival and adaptation. Altogether, our findings provide evidence for the biological role of the TNT-mediated vesicle exchange between stromal and leukemic cells, implicating the direct vesicle and protein transfer in the stroma-provided protection of leukemic cells.

Genomes of Neutrophilic Sulfur-Oxidizing Chemolithoautotrophs Representing 9 Proteobacterial Species From 8 Genera

Even in the current era of metagenomics, the interpretation of nucleotide sequence data is primarily dependent on knowledge obtained from a limited number of microbes isolated in pure culture. Thus, it is of fundamental importance to expand the variety of strains available in pure culture, to make reliable connections between physiological characteristics and genomic information. In this study, two sulfur oxidizers that potentially represent two novel species were isolated and characterized. They were subjected to whole-genome sequencing together with 7 neutrophilic and chemolithoautotrophic sulfur-oxidizing bacteria. The genes for sulfur oxidation in the obtained genomes were identified and compared with those of isolated sulfur oxidizers in the classes Betaproteobacteria and Gammaproteobacteria. Although the combinations of these genes in the respective genomes are diverse, typical combinations corresponding to three types of core sulfur oxidation pathways were identified. Each pathway involves one of three specific sets of proteins, SoxCD, DsrABEFHCMKJOP, and HdrCBAHypHdrCB. All three core pathways contain the SoxXYZAB proteins, and a cytoplasmic sulfite oxidase encoded by soeABC is a conserved component in the core pathways lacking SoxCD. Phylogenetically close organisms share same core sulfur oxidation pathway, but a notable exception was observed in the family ‘Sulfuricellaceae’. In this family, some strains have either core pathway involving DsrABEFHCMKJOP or HdrCBAHypHdrCB, while others have both pathways. A proteomics analysis showed that proteins constituting the core pathways were produced at high levels. While hypothesized function of HdrCBAHypHdrCB is similar to that of Dsr system, both sets of proteins were detected with high relative abundances in the proteome of a strain possessing genes for these proteins. In addition to the genes for sulfur oxidation, those for arsenic metabolism were searched for in the sequenced genomes. As a result, two strains belonging to the families Thiobacillaceae and Sterolibacteriaceae were observed to harbor genes encoding ArxAB, a type of arsenite oxidase that has been identified in a limited number of bacteria. These findings were made with the newly obtained genomes, including those from 6 genera from which no genome sequence of an isolated organism was previously available. These genomes will serve as valuable references to interpret nucleotide sequences.

Dynamics and functions of lipid droplets.

Lipid droplets are storage organelles at the centre of lipid and energy homeostasis. They have a unique architecture consisting of a hydrophobic core of neutral lipids, which is enclosed by a phospholipid monolayer that is decorated by a specific set of proteins. Originating from the endoplasmic reticulum, lipid droplets can associate with most other cellular organelles through membrane contact sites. It is becoming apparent that these contacts between lipid droplets and other organelles are highly dynamic and coupled to the cycles of lipid droplet expansion and shrinkage. Importantly, lipid droplet biogenesis and degradation, as well as their interactions with other organelles, are tightly coupled to cellular metabolism and are critical to buffer the levels of toxic lipid species. Thus, lipid droplets facilitate the coordination and communication between different organelles and act as vital hubs of cellular metabolism.

Proteomic analysis of exosomes reveals an association between cell invasiveness and exosomal bioactivity on endothelial and mesenchymal cell migration in vitro.

Ovarian cancer has resulted in over 140 000 deaths reported annually worldwide. This is often attributed to cellular changes in the microenvironment, including increased migration of mesenchymal stem cells (MSCs) and endothelial cells (ECs) to facilitate metastasis. Recently, the ability of exosomes to communicate signals between cells (and promote cancer progression) has been established. In the present study, we explored the effect of exosomes on cells present in the tumour microenvironment. Exosomes were isolated from ovarian cancer cells with different invasive capacity (high = SKOV-3 and low = OVCAR-3) by differential and buoyant density centrifugation and characterised using nanoparticle tracking analysis (NTA), Western blot, and EM. Exosome secretion was positively correlated with invasiveness of releasing cells. Proteomic analyses identified common and unique proteins between exosomes from SKOV-3 and OVCAR-3 with gene ontology analyses revealing that these exosomes are involved in the regulation of cell migration. Since the tumour microenvironment contains multiple cell types, including MSCs and ECs, we examined the effect of these exosomes on MSC and EC migration.Exosomes promoted MSC and EC migration in a time- and concentration-dependent manner. The effect of exosomes isolated from SKOV-3 on cell migration was significantly higher compared with exosomes from OVCAR-3. Thus, we suggest that exosomes from ovarian cancer cells contain a specific set of proteins that are representative of its cell of origin and the invasive capacity.

P-Body Localization of the Hrr25/Casein Kinase 1 Protein Kinase Is Required for the Completion of Meiosis

P-bodies are liquid droplet-like compartments that lack a limiting membrane and are present in many eukaryotic cells. These structures contain specific sets of proteins and mRNAs at concentrations higher than that in the surrounding environment. Although highly conserved, the normal physiological roles of these ribonucleoprotein (RNP) granules remain poorly defined. Here, we report that P-bodies are required for the efficient completion of meiosis in the budding yeast Saccharomyces cerevisiae P-bodies were found to be present during all phases of the meiotic program and to provide protection for the Hrr25/CK1 protein kinase, a key regulator of this developmental process. A failure to associate with these RNP granules resulted in diminished levels of Hrr25 and an ensuing inability to complete meiosis. This work therefore identifies a novel function for these RNP granules and indicates how protein recruitment to these structures can have a significant impact on eukaryotic cell biology.