JOB RESPONSIBILITIES Identify quality oriented process improvement initiatives and technological advancements; identify, plan, lead, and execute research and development activities across all Graniterock products lines including but not limited to Cold-In-Place (CIR) Recycling, Hot Mix Asphalt,

A team of researchers based in London, Ont. is the first in the world to attempt treating critical COVID-19 patients with a modified form of dialysis, doctors say.

PENSIONERS spend just five years of their retirement in good health, warns new research.

As the all-out effort for a vaccine accelerates, USA TODAY is rounding up some of the week's most notable developments.

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Life changed for Avon High School alum Jesse Kramer in June, when he made his TV network debut on "The Four: Battle for Stardom."

Indianapolis Prize finalist Christophe Boesch has spent half a century researching chimpanzees and their behaviors.

Provincial police have located human remains in their search for a missing Meaford man, and one person has been taken into custody.

A team of researchers at the University of California at San Diego have created a soft robotic lens that responds to eye movements.

Japanese researchers at Keio University have unveiled a robotic tail that has been designed to be worn by elderly people who struggle to maintain their balance.

The "Alias" actor was diagnosed with the disease as a child.

There are several things which go into successful video marketing on YouTube. One of these things is the very often neglected YouTube SEO that’s so vital for getting viewers to actually find your video. Too many YouTube marketers are concerned with what’s going on in their videos to really push for better SEO, and it […]

The post How Web Media Developer can use YouTube SEO with Better Search Marketing to Rank Their Videos appeared first on SpyreStudios.

Keyword research is the backbone of your online presence. It’s common knowledge that this practice determines your rankings and visibility in the organic search. However, the way keyword research is done has changed over time. Today, using Google’s Keyword Planner to find phrases with a high search volume and optimizing your site for them is not […]

The post How to do Keyword Research for SEO appeared first on ReadWrite.

Local news publishers have struggled to monetize spike in consumer interest.

Please visit Search Engine Land for the full article.

The Rich Results Test tool now also supports SpecialAnnouncement markup.

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‘The idea that what’s happening with searcher behavior is not causing these shifts means that Google is in there writing that code for every intent, every day, and I can’t believe that’s what’s happening,’ said Dr. Pete Meyers on Live with Search Engine Land.

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If you are having indexing issues with your HTML or JavaScript-based website then you won’t want to miss this chat.

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Even the biggest sites on the internet can have problems with their SEO and Google.

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The suggestions surface for multiple intents for logged-in users.

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This is the first time Google says it has provided this kind of data to the public.

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This update was an “absolute monster” said Mordy Oberstein from RankRanger.

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As the coronavirus crisis built to pandemic levels in early March, a relatively unknown tech company confronted a defining opportunity. Zoom Video Communications, a fast-growing enterprise videoconferencing platform with roots in both Silicon Valley and China, had already seen its market cap grow from under $10 billion to nearly double that. As the coronavirus began … Continue reading "New Research Shows Why and How Zoom Could Become an Advertising Driven Business"

The Delivery Man Synopsis The Delivery Man is a comedy television series produced by ITV that premiered in April of 2015. The Delivery Man follows the story of Matthew (played by Darren Boyd – Spy), a police officer who decides to switch careers in his mid-thirties and becomes a midwife. The Delivery Man is currently […]

Other Space Synopsis Other Space is a Science Fiction comedy television series produced by Yahoo that premiered in April of 2015. Created by Paul Feig (Freaks and Geeks) Other Space is set in the early 22nd century and follows the story of the inexperienced crew aboard UMP Cruiser on a routine space exploratory mission. In […]

Peter Kay’s Car Share Synopsis Peter Kay’s Car Share is a situation comedy television series produced by BBC One that premiered in April of 2015. Peter Kay’s Car Share follows the story of John (played by Peter Kay), an assistant manager in a supermarket and his co-worker and promotions rep Kayleigh (played by Sian Gibson). […]

Meet the Smiths Synopsis Meet the Smiths is a scripted reality television series produced by TBS that premiered in April of 2015. Meet the Smiths follows the daily lives of Kenny Smith, an NBA analyst for TNT and his wife Gwendolyn Osborne-Smith, a model for The Price is Right. The couple has five children together […]

Arranged Synopsis Arranged is a reality television series produced by the FYI Network that premiered in April of 2015. Arranged follows the story of the three American couples who all come from different backgrounds, but their cultural traditions allow for arranged marriages. The series chronicles their lives in the lead up to the wedding and […]

Wayward Pines Synopsis Wayward Pines is a mystery thriller television series produced by Fox Network and set to premiere in May of 2015. Based on the book Pines written by Blake Crouch and produced by M. Night Shyamalan Wayward Pines follows the story of an FBI agent Ethan Bruke (played by Matt Dillon), who travels […]

Miles from Tomorrowland Synopsis Miles from Tomorrowland is an animated television series produced by the Disney Channel that premiered in February of 2015. Miles from Tomorrowland is an American animated TV series about the adventures of an inquisitive, if somewhat reckless boy named Miles who travels the universe with his family. Miles from Tomorrowland is […]

The Protectors (1972) Synopsis The Protectors (1972) was an action thriller television series produced by ITV that premiered in September of 1972. The Protectors (1972) was a British action thriller series created by Gerry Anderson (Thunderbirds) about an international group of rich troubleshooters. Contessa di Contini (played by Nyree Dawn Porter), Paul Buchet (played by […]

Wormwood Synopsis Wormwood was a children’s television series produced by Network Ten and ABC1 that premiered in October of 2007. Made as a joint Canadian-Australian production, Wormwood was a TV series about a quirky town where a group of kids investigated strange events. The series was based on the stories written by Paul Jennings. Wormwood […]

A Sharp Intake Of Breath Synopsis A Sharp Intake Of Breath was a situation comedy television series produced by ITV that premiered in February of 1978. Created by Ronnie Taylor A Sharp Intake Of Breath followed the story of Peter Barnes (played by David Jason – Only Fools And Horses), an ordinary man who made […]

An eight-year-old boy with good intentions sparked a missing person search on Tuesday.

Grey Bruce provincial police say the OPP canine unit found human remains in Meaford, and one person is under arrest.

The following article, Univ of Pittsburg Researcher, China Native, Dr. Bing Liu, “On the verge” of COVID19 Breakthrough Is Murdered…Alleged Gunman, Hao Gu, Kills Himself, was first published on 100PercentFedUp.com.

A 37-year-old China native and "outstanding researcher" at the University of Pittsburgh...

Continue reading: Univ of Pittsburg Researcher, China Native, Dr. Bing Liu, “On the verge” of COVID19 Breakthrough Is Murdered…Alleged Gunman, Hao Gu, Kills Himself ...

Chatham House is pleased to announce that Professor Tim Benton has been appointed as research director of the Energy, Environment and Resources Department.

6 April 2020

Ignat V. Shilov
Sep 1, 2007; 6:1638-1655
Technology

Invitation Only Research Event

Event participants

Rebecca Wolfe, Lecturer, Harris School for Public Policy and Associate, Pearson Institute for the Study and Resolution of Global Conflicts, University of Chicago
Tom Gillhespy, Principal Consultant, Itad
Shodmon Hojibekov, Chief Executive Officer, Aga Khan Agency for Habitat (Afghanistan)
Chair: Champa Patel, Director, Asia-Pacific Programme, Chatham House

6 April 2020

Invitation Only Research Event

Event participants

Rebecca Wolfe, Lecturer, Harris School for Public Policy and Associate, Pearson Institute for the Study and Resolution of Global Conflicts, University of Chicago
Tom Gillhespy, Principal Consultant, Itad
Shodmon Hojibekov, Chief Executive Officer, Aga Khan Agency for Habitat (Afghanistan)
Chair: Champa Patel, Director, Asia-Pacific Programme, Chatham House

Systemic infection and proliferation of intracellular pathogens require the biogenesis of a growth-stimulating compartment. The gastrointestinal pathogen Salmonella enterica commonly forms highly dynamic and extensive tubular membrane compartments built from Salmonella-modified membranes (SMMs) in diverse host cells. Although the general mechanism involved in the formation of replication-permissive compartments of S. enterica is well researched, much less is known regarding specific adaptations to different host cell types. Using an affinity-based proteome approach, we explored the composition of SMMs in murine macrophages. The systematic characterization provides a broader landscape of host players to the maturation of Salmonella-containing compartments and reveals core host elements targeted by Salmonella in macrophages as well as epithelial cells. However, we also identified subtle host specific adaptations. Some of these observations, such as the differential involvement of the COPII system, Rab GTPases 2A, 8B, 11 and ER transport proteins Sec61 and Sec22B may explain cell line-dependent variations in the pathophysiology of Salmonella infections. In summary, our system-wide approach demonstrates a hitherto underappreciated impact of the host cell type in the formation of intracellular compartments by Salmonella.

Acute myeloid leukemia (AML) is a clonal disorder arising from hematopoietic myeloid progenitors. Aberrantly activated tyrosine kinases (TK) are involved in leukemogenesis and are associated with poor treatment outcome. Kinase inhibitor (KI) treatment has shown promise in improving patient outcome in AML. However, inhibitor selection for patients is suboptimal.

In a preclinical effort to address KI selection, we analyzed a panel of 16 AML cell lines using phosphotyrosine (pY) enrichment-based, label-free phosphoproteomics. The Integrative Inferred Kinase Activity (INKA) algorithm was used to identify hyperphosphorylated, active kinases as candidates for KI treatment, and efficacy of selected KIs was tested.

Heterogeneous signaling was observed with between 241 and 2764 phosphopeptides detected per cell line. Of 4853 identified phosphopeptides with 4229 phosphosites, 4459 phosphopeptides (4430 pY) were linked to 3605 class I sites (3525 pY). INKA analysis in single cell lines successfully pinpointed driver kinases (PDGFRA, JAK2, KIT and FLT3) corresponding with activating mutations present in these cell lines. Furthermore, potential receptor tyrosine kinase (RTK) drivers, undetected by standard molecular analyses, were identified in four cell lines (FGFR1 in KG-1 and KG-1a, PDGFRA in Kasumi-3, and FLT3 in MM6). These cell lines proved highly sensitive to specific KIs. Six AML cell lines without a clear RTK driver showed evidence of MAPK1/3 activation, indicative of the presence of activating upstream RAS mutations. Importantly, FLT3 phosphorylation was demonstrated in two clinical AML samples with a FLT3 internal tandem duplication (ITD) mutation.

Our data show the potential of pY-phosphoproteomics and INKA analysis to provide insight in AML TK signaling and identify hyperactive kinases as potential targets for treatment in AML cell lines. These results warrant future investigation of clinical samples to further our understanding of TK phosphorylation in relation to clinical response in the individual patient.

Human leukocyte antigen (HLA) B*51:01 and endoplasmic reticulum aminopeptidase 1 (ERAP1) are strongly genetically associated with Behcet's disease (BD). Previous studies have defined two subgroups of HLA-B*51 peptidome containing proline (Pro) or alanine (Ala) at position 2 (P2). Little is known about the unconventional non-Pro/Ala2 HLA-B*51-bound peptides. We aimed to study the features of this novel subpeptidome, and investigate its regulation by ERAP1. CRISPR-Cas9 was used to generate an HLA-ABC-triple knockout HeLa cell line (HeLa.ABC-KO), which was subsequently transduced to express HLA-B*51:01 (HeLa.ABC-KO.B51). ERAP1 was silenced using lentiviral shRNA. Peptides bound to HLA-B*51:01 were eluted and analyzed by mass spectrometry. The characteristics of non-Pro/Ala2, Pro2, and Ala2 peptides and their alteration by ERAP1 silencing were investigated. Effects of ERAP1 silencing on cell surface expression of HLA-B*51:01 were studied using flow cytometry. More than 20% of peptides eluted from HLA-B*51:01 lacked Pro or Ala at P2. This unconventional group of HLA-B*51:01-bound peptides was relatively enriched for 8-mers (with relatively fewer 9-mers) compared with the Pro2 and Ala2 subpeptidomes and had similar N-terminal and C-terminal residue usages to Ala2 peptides (with the exception of the less abundant leucine at position ). Knockdown of ERAP1 increased the percentage of non-Pro/Ala2 from 20% to ~40%, increased the percentage of longer (10-mer and 11-mer) peptides eluted from HLA-B*51:01 complexes, and abrogated the predominance of leucine at P1. Interestingly knockdown of ERAP1 altered the length and N-terminal residue usage of non-Ala2&Pro2 and Ala2 but not the Pro2 peptides. Finally, ERAP1 silencing regulated the expression levels of cell surface HLA-B*51 in a cell-type-dependent manner. In conclusion, we have used a novel methodology to identify an unconventional but surprisingly abundant non-Pro/Ala2 HLA-B*51:01 subpeptidome. It is increased by knockdown of ERAP1, a gene affecting the risk of developing BD. This has implications for theories of disease pathogenesis.

The redox-based modifications of cysteine residues in proteins regulate their function in many biological processes. The gas molecule H₂S has been shown to persulfidate redox sensitive cysteine residues resulting in an H₂S-modified proteome known as the sulfhydrome. Tandem Mass Tags (TMT) multiplexing strategies for large-scale proteomic analyses have become increasingly prevalent in detecting cysteine modifications. Here we developed a TMT-based proteomics approach for selectively trapping and tagging cysteine persulfides in the cellular proteomes. We revealed the natural protein sulfhydrome of two human cell lines, and identified insulin as a novel substrate in pancreatic beta cells. Moreover, we showed that under oxidative stress conditions, increased H₂S can target enzymes involved in energy metabolism by switching specific cysteine modifications to persulfides. Specifically, we discovered a Redox Thiol Switch, from protein S-glutathioinylation to S-persulfidation (RTS^GS). We propose that the RTS^GS from S-glutathioinylation to S-persulfidation is a potential mechanism to fine tune cellular energy metabolism in response to different levels of oxidative stress.

Laser-capture microdissection (LCM) allows the visualization and isolation of morphologically distinct subpopulations of cells from heterogeneous tissue specimens. In combination with formalin-fixed and paraffin-embedded (FFPE) tissue it provides a powerful tool for retrospective and clinically relevant studies of tissue proteins in a healthy and diseased context. We first optimized the protocol for efficient LCM analysis of FFPE tissue specimens. The use of SDS containing extraction buffer in combination with the single-pot solid-phase-enhanced sample preparation (SP3) digest method gave the best results regarding protein yield and protein/peptide identifications. Microdissected FFPE human substantia nigra tissue samples (~3,000 cells) were then analyzed, using tandem mass tag (TMT) labeling and LC-MS/MS, resulting in the quantification of >5,600 protein groups. Nigral proteins were classified and analyzed by abundance, showing an enrichment of extracellular exosome and neuron-specific gene ontology (GO) terms among the higher abundance proteins. Comparison of microdissected samples with intact tissue sections, using a label-free shotgun approach, revealed an enrichment of neuronal cell type markers, such as tyrosine hydroxylase and alpha-synuclein, as well as proteins annotated with neuron-specific GO terms. Overall, this study provides a detailed protocol for laser-capture proteomics using FFPE tissue and demonstrates the efficiency of LCM analysis of distinct cell subpopulations for proteomic analysis using low sample amounts.

Mass spectrometry (MS)-based proteomics has great potential for overcoming the limitations of antibody-based immunoassays for antibody-independent, comprehensive, and quantitative proteomic analysis of single cells. Indeed, recent advances in nanoscale sample preparation have enabled effective processing of single cells. In particular, the concept of using boosting/carrier channels in isobaric labeling to increase the sensitivity in MS detection has also been increasingly used for quantitative proteomic analysis of small-sized samples including single cells. However, the full potential of such boosting/carrier approaches has not been significantly explored, nor has the resulting quantitation quality been carefully evaluated. Herein, we have further evaluated and optimized our recent boosting to amplify signal with isobaric labeling (BASIL) approach, originally developed for quantifying phosphorylation in small number of cells, for highly effective analysis of proteins in single cells. This improved BASIL (iBASIL) approach enables reliable quantitative single-cell proteomics analysis with greater proteome coverage by carefully controlling the boosting-to-sample ratio (e.g. in general <100x) and optimizing MS automatic gain control (AGC) and ion injection time settings in MS/MS analysis (e.g. 5E5 and 300 ms, respectively, which is significantly higher than that used in typical bulk analysis). By coupling with a nanodroplet-based single cell preparation (nanoPOTS) platform, iBASIL enabled identification of ~2500 proteins and precise quantification of ~1500 proteins in the analysis of 104 FACS-isolated single cells, with the resulting protein profiles robustly clustering the cells from three different acute myeloid leukemia cell lines. This study highlights the importance of carefully evaluating and optimizing the boosting ratios and MS data acquisition conditions for achieving robust, comprehensive proteomic analysis of single cells.

HNF4α is a nuclear receptor produced as 12 isoforms from two promoters by alternative splicing. To characterize the transcriptional capacities of all 12 HNF4α isoforms, stable lines expressing each isoform were generated. The entire transcriptome associated with each isoform was analyzed as well as their respective interacting proteome. Major differences were noted in the transcriptional function of these isoforms. The α1 and α2 isoforms were the strongest regulators of gene expression whereas the α3 isoform exhibited significantly reduced activity. The α4, α5, and α6 isoforms, which use an alternative first exon, were characterized for the first time, and showed a greatly reduced transcriptional potential with an inability to recognize the consensus response element of HNF4α. Several transcription factors and coregulators were identified as potential specific partners for certain HNF4α isoforms. An analysis integrating the vast amount of omics data enabled the identification of transcriptional regulatory mechanisms specific to certain HNF4α isoforms, hence demonstrating the importance of considering all isoforms given their seemingly diverse functions.

The respiratory epithelium comprises polarized cells at the interface between the environment and airway tissues. Polarized apical and basolateral protein secretions are a feature of airway epithelium homeostasis. Human respiratory syncytial virus (hRSV) is a major human pathogen that primarily targets the respiratory epithelium. However, the consequences of hRSV infection on epithelium secretome polarity and content remain poorly understood. To investigate the hRSV-associated apical and basolateral secretomes, a proteomics approach was combined with an ex vivo pediatric human airway epithelial (HAE) model of hRSV infection (data are available via ProteomeXchange and can be accessed at https://www.ebi.ac.uk/pride/ with identifier PXD013661). Following infection, a skewing of apical/basolateral abundance ratios was identified for several individual proteins. Novel modulators of neutrophil and lymphocyte activation (CXCL6, CSF3, SECTM1 or CXCL16), and antiviral proteins (BST2 or CEACAM1) were detected in infected, but not in uninfected cultures. Importantly, CXCL6, CXCL16, CSF3 were also detected in nasopharyngeal aspirates (NPA) from hRSV-infected infants but not healthy controls. Furthermore, the antiviral activity of CEACAM1 against RSV was confirmed in vitro using BEAS-2B cells. hRSV infection disrupted the polarity of the pediatric respiratory epithelial secretome and was associated with immune modulating proteins (CXCL6, CXCL16, CSF3) never linked with this virus before. In addition, the antiviral activity of CEACAM1 against hRSV had also never been previously characterized. This study, therefore, provides novel insights into RSV pathogenesis and endogenous antiviral responses in pediatric airway epithelium.