The Comparison of the Effects of Nodular Cast Iron Laser Alloying with Selected Substances.

The aim of this research was to compare the effects of laser treatment, with the same heating conditions, using four selected alloying substances (silicon, cobalt, silicon nitride and titanium), in the surface layer of nodular cast iron. The treatment was performed with a molecular laser. As the microstructure observation revealed, the greatest amount of implemented elements was diluted during the treatment in a solid solution. In all cases (except during the alloying process with cobalt), in the alloying zone, a fine and homogeneous microstructure was found. In the alloying zone, cobalt counteracted the formation of the martensitic microstructure so effectively that austenite turned into exclusively fine perlite (or bainite at most). The size of the obtained alloyed zone was different, despite the same laser heat treatment parameters. A 30% smaller depth of zone after laser alloying with silicon nitride, as compared with alloying with cobalt or silicon, was observed. The highest strengthening of the alloyed zone could be expected when silicon (hardness was approx. 980HV0.1 and the modulus of elasticity was 208 GPa) and titanium (hardness was approx. 880HV0.1 and the modulus of elasticity was 194 GPa) were used. The lowest hardness (700HV0.1) was observed for the zone alloyed with cobalt due to pearlite (or bainite) existence.

Microstructure and Soil Wear Resistance of a Grey Cast Iron Alloy Reinforced with Ni and Cr Laser Coatings.

The goal of the presented investigation was to assess the impact of surface laser modification with the implementation of nickel and chromium on the microstructure and tribological behaviour of grey iron. Surface laser modification consisted of remelting the surface layer with simultaneous implementation of selected elements. In the first variant of treatment only nickel was implemented and in the second one, a combination of nickel with chromium together. This treatment was performed on an agriculture machine part made of grey iron and working in intensive friction conditions. The constituted surface layer was characterized by about 0.45 mm of depth and a 160 mm2 area of the most exposed to wear of the treated part. In the case of both types of variants, the achieved surface layer microstructure was identified as homogenized with small grains. It involved nickel in the first variant of modification and nickel and chromium in the second one. The attained microstructure with nickel addition was characterized by nearly 800 HV0.1 of hardness (a 3.6-fold increase in comparison to its core material). The approximate hardness of 900 HV0.1 was achieved in the case of the microstructure enriched with nickel and chromium (over a 4-fold increase in comparison to the core material). The roughness of the surface after laser modification was reduced (nearly 3-fold) in comparison to the original surface of the part that was characterized by quite substantial coarseness. The wear test showed that Ni and Cr laser coatings increased resistance to abrasive wear resulting from the modification of the microstructure by the formation of martensite and grain fragmentation. Laser modified parts had a 2.5-fold smaller mass loss than untreated parts. Both types of performed variants: with the implementation of nickel and a combination of nickel and chromium gave comparable effects.

Reliability Analysis for Unrepairable Automotive Components.

The analysis of the reliability parameters of a technical object and the determination of the change in the reliability of the object over time, requires the knowledge of the functional characteristics and reliability parameters of the elements included in a system. On the basis of the failure data of the selected element of the object, in this case the vehicle, it is possible to determine the average working time to failure of the element and the appropriate form of distribution that characterizes the reliability and durability parameters of the tested element. The main purpose of the research presented in the article was to develop a method of assessing the reliability of an electronic component of a vehicle-a boot lid contactor. This paper also presents three possible methods of repairing the boot lid contactor (sealing the housing with adhesive with better way, replacing the element with a new one or the most time-consuming solution, changing the shape of the boot lid). The authors also decided to determine the reliability and cost parameters that will allow preventive replacement of this element. The tests were carried out on a fleet of 61 vehicles of the same model, but with different body structures. Contactor failures were reported in 41 cases, of which 29 were in the hatchback construction and 12 in the estate type. The analysis of the distribution selection for the tested part of the passenger car-the boot lid contactor-was performed using the Likelihood Value (LKV) test to determine the rank of distributions. Also the maximum likelihood (MLE) method was used to estimate the distribution parameters. The three-parameter Weibull distribution was the best-fitted distribution in both cases. It was clearly defined that one model of car with two different types of body have vastly different reliability characteristic. Based on the reliability characteristic and parameters, the appropriate preventive actions can be taken, minimizing the risk of damage, thus avoiding financial losses and guaranteeing an appropriate level of vehicle safety.

Integrative pathway enrichment analysis of multivariate omics data.

Multi-omics datasets represent distinct aspects of the central dogma of molecular biology. Such high-dimensional molecular profiles pose challenges to data interpretation and hypothesis generation. ActivePathways is an integrative method that discovers significantly enriched pathways across multiple datasets using statistical data fusion, rationalizes contributing evidence and highlights associated genes. As part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2658 cancers across 38 tumor types, we integrated genes with coding and non-coding mutations and revealed frequently mutated pathways and additional cancer genes with infrequent mutations. We also analyzed prognostic molecular pathways by integrating genomic and transcriptomic features of 1780 breast cancers and highlighted associations with immune response and anti-apoptotic signaling. Integration of ChIP-seq and RNA-seq data for master regulators of the Hippo pathway across normal human tissues identified processes of tissue regeneration and stem cell regulation. ActivePathways is a versatile method that improves systems-level understanding of cellular organization in health and disease through integration of multiple molecular datasets and pathway annotations.

Pathway and network analysis of more than 2500 whole cancer genomes.

The catalog of cancer driver mutations in protein-coding genes has greatly expanded in the past decade. However, non-coding cancer driver mutations are less well-characterized and only a handful of recurrent non-coding mutations, most notably TERT promoter mutations, have been reported. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2658 cancer across 38 tumor types, we perform multi-faceted pathway and network analyses of non-coding mutations across 2583 whole cancer genomes from 27 tumor types compiled by the ICGC/TCGA PCAWG project that was motivated by the success of pathway and network analyses in prioritizing rare mutations in protein-coding genes. While few non-coding genomic elements are recurrently mutated in this cohort, we identify 93 genes harboring non-coding mutations that cluster into several modules of interacting proteins. Among these are promoter mutations associated with reduced mRNA expression in TP53, TLE4, and TCF4. We find that biological processes had variable proportions of coding and non-coding mutations, with chromatin remodeling and proliferation pathways altered primarily by coding mutations, while developmental pathways, including Wnt and Notch, altered by both coding and non-coding mutations. RNA splicing is primarily altered by non-coding mutations in this cohort, and samples containing non-coding mutations in well-known RNA splicing factors exhibit similar gene expression signatures as samples with coding mutations in these genes. These analyses contribute a new repertoire of possible cancer genes and mechanisms that are altered by non-coding mutations and offer insights into additional cancer vulnerabilities that can be investigated for potential therapeutic treatments.

Candidate Cancer Driver Mutations in Distal Regulatory Elements and Long-Range Chromatin Interaction Networks.

A comprehensive catalog of cancer driver mutations is essential for understanding tumorigenesis and developing therapies. Exome-sequencing studies have mapped many protein-coding drivers, yet few non-coding drivers are known because genome-wide discovery is challenging. We developed a driver discovery method, ActiveDriverWGS, and analyzed 120,788 cis-regulatory modules (CRMs) across 1,844 whole tumor genomes from the ICGC-TCGA PCAWG project. We found 30 CRMs with enriched SNVs and indels (FDR < 0.05). These frequently mutated regulatory elements (FMREs) were ubiquitously active in human tissues, showed long-range chromatin interactions and mRNA abundance associations with target genes, and were enriched in motif-rewiring mutations and structural variants. Genomic deletion of one FMRE in human cells caused proliferative deficiencies and transcriptional deregulation of cancer genes CCNB1IP1, CDH1, and CDKN2B, validating observations in FMRE-mutated tumors. Pathway analysis revealed further sub-significant FMREs at cancer genes and processes, indicating an unexplored landscape of infrequent driver mutations in the non-coding genome.

Global phosphoproteomic analysis identifies SRMS-regulated secondary signaling intermediates.

BACKGROUND: The non-receptor tyrosine kinase, SRMS (Src-related kinase lacking C-terminal regulatory tyrosine and N-terminal myristoylation sites) is a member of the BRK family kinases (BFKs) which represents an evolutionarily conserved relative of the Src family kinases (SFKs). Tyrosine kinases are known to regulate a number of cellular processes and pathways via phosphorylating substrate proteins directly and/or by partaking in signaling cross-talks leading to the indirect modulation of various signaling intermediates. In a previous study, we profiled the tyrosine-phosphoproteome of SRMS and identified multiple candidate substrates of the kinase. The broader cellular signaling intermediates of SRMS are unknown. METHODS: In order to uncover the broader SRMS-regulated phosphoproteome and identify the SRMS-regulated indirect signaling intermediates, we performed label-free global phosphoproteomics analysis on cells expressing wild-type SRMS. Using computational database searching and bioinformatics analyses we characterized the dataset. RESULTS: Our analyses identified 60 hyperphosphorylated (phosphoserine/phosphothreonine) proteins mapped from 140 hyperphosphorylated peptides. Bioinfomatics analyses identified a number of significantly enriched biological and cellular processes among which DNA repair pathways were found to be upregulated while apoptotic pathways were found to be downregulated. Analyses of motifs derived from the upregulated phosphosites identified Casein kinase 2 alpha (CK2α) as one of the major potential kinases contributing to the SRMS-dependent indirect regulation of signaling intermediates. CONCLUSIONS: Overall, our phosphoproteomics analyses identified serine/threonine phosphorylation dynamics as important secondary events of the SRMS-regulated phosphoproteome with implications in the regulation of cellular and biological processes.

Pathogenic Germline Variants in 10,389 Adult Cancers.

We conducted the largest investigation of predisposition variants in cancer to date, discovering 853 pathogenic or likely pathogenic variants in 8% of 10,389 cases from 33 cancer types. Twenty-one genes showed single or cross-cancer associations, including novel associations of SDHA in melanoma and PALB2 in stomach adenocarcinoma. The 659 predisposition variants and 18 additional large deletions in tumor suppressors, including ATM, BRCA1, and NF1, showed low gene expression and frequent (43%) loss of heterozygosity or biallelic two-hit events. We also discovered 33 such variants in oncogenes, including missenses in MET, RET, and PTPN11 associated with high gene expression. We nominated 47 additional predisposition variants from prioritized VUSs supported by multiple evidences involving case-control frequency, loss of heterozygosity, expression effect, and co-localization with mutations and modified residues. Our integrative approach links rare predisposition variants to functional consequences, informing future guidelines of variant classification and germline genetic testing in cancer.

Phosphoproteomics Analysis Identifies Novel Candidate Substrates of the Nonreceptor Tyrosine Kinase, Src-related Kinase Lacking C-terminal Regulatory Tyrosine and N-terminal Myristoylation Sites (SRMS).

SRMS (Src-related kinase lacking C-terminal regulatory tyrosine and N-terminal myristoylation sites), also known as PTK 70 (Protein tyrosine kinase 70), is a non-receptor tyrosine kinase that belongs to the BRK family of kinases (BFKs). To date less is known about the cellular role of SRMS primarily because of the unidentified substrates or signaling intermediates regulated by the kinase. In this study, we used phosphotyrosine antibody-based immunoaffinity purification in large-scale label-free quantitative phosphoproteomics to identify novel candidate substrates of SRMS. Our analyses led to the identification of 1258 tyrosine-phosphorylated peptides which mapped to 663 phosphoproteins, exclusively from SRMS-expressing cells. DOK1, a previously characterized SRMS substrate, was also identified in our analyses. Functional enrichment analyses revealed that the candidate SRMS substrates were enriched in various biological processes including protein ubiquitination, mitotic cell cycle, energy metabolism and RNA processing, as well as Wnt and TNF signaling. Analyses of the sequence surrounding the phospho-sites in these proteins revealed novel candidate SRMS consensus substrate motifs. We utilized customized high-throughput peptide arrays to validate a subset of the candidate SRMS substrates identified in our MS-based analyses. Finally, we independently validated Vimentin and Sam68, as bona fide SRMS substrates through in vitro and in vivo assays. Overall, our study identified a number of novel and biologically relevant SRMS candidate substrates, which suggests the involvement of the kinase in a vast array of unexplored cellular functions.

ActiveDriverDB: human disease mutations and genome variation in post-translational modification sites of proteins.

Interpretation of genetic variation is needed for deciphering genotype-phenotype associations, mechanisms of inherited disease, and cancer driver mutations. Millions of single nucleotide variants (SNVs) in human genomes are known and thousands are associated with disease. An estimated 21% of disease-associated amino acid substitutions corresponding to missense SNVs are located in protein sites of post-translational modifications (PTMs), chemical modifications of amino acids that extend protein function. ActiveDriverDB is a comprehensive human proteo-genomics database that annotates disease mutations and population variants through the lens of PTMs. We integrated >385,000 published PTM sites with ∼3.6 million substitutions from The Cancer Genome Atlas (TCGA), the ClinVar database of disease genes, and human genome sequencing projects. The database includes site-specific interaction networks of proteins, upstream enzymes such as kinases, and drugs targeting these enzymes. We also predicted network-rewiring impact of mutations by analyzing gains and losses of kinase-bound sequence motifs. ActiveDriverDB provides detailed visualization, filtering, browsing and searching options for studying PTM-associated mutations. Users can upload mutation datasets interactively and use our application programming interface in pipelines. Integrative analysis of mutations and PTMs may help decipher molecular mechanisms of phenotypes and disease, as exemplified by case studies of TP53, BRCA2 and VHL. The open-source database is available at https://www.ActiveDriverDB.org.

Protein-peptide molecular docking with large-scale conformational changes: the p53-MDM2 interaction.

Protein-peptide interactions are often associated with large-scale conformational changes that are difficult to study either by classical molecular modeling or by experiment. Recently, we have developed the CABS-dock method for flexible protein-peptide docking that enables large-scale rearrangements of the protein chain. In this study, we use CABS-dock to investigate the binding of the p53-MDM2 complex, an element of the cell cycle regulation system crucial for anti-cancer drug design. Experimental data suggest that p53-MDM2 binding is affected by significant rearrangements of a lid region - the N-terminal highly flexible MDM2 fragment; however, the details are not clear. The large size of the highly flexible MDM2 fragments makes p53-MDM2 intractable for exhaustive binding dynamics studies using atomistic models. We performed extensive dynamics simulations using the CABS-dock method, including large-scale structural rearrangements of MDM2 flexible regions. Without a priori knowledge of the p53 peptide structure or its binding site, we obtained near-native models of the p53-MDM2 complex. The simulation results match well the experimental data and provide new insights into the possible role of the lid fragment in p53 binding. The presented case study demonstrates that CABS-dock methodology opens up new opportunities for protein-peptide docking with large-scale changes of the protein receptor structure.

Decaying mouse volatiles perceived by Calliphora vicina Rob.-Desv.

Volatiles emitted by decaying human remains are in the focus of recent research. The identification of core volatiles in this field is of high importance, because cadaveric volatiles generally show high variation. In this study, the volatile profiles of five mice (Myodes glareolus) were sampled with charcoal filter tubes from their time of death until advanced decay. Eleven compounds were quantitated by means of gas chromatography-mass spectrometry. Electroantennographic experiments with female Calliphora vicina antennae led to the identification of dimethyl trisulfide, dimethyl disulfide, nonanal, hexan-1-ol, 1-octen-3-ol, 3-methylbutan-1-ol, and heptanal as electrophysiologically active compounds. When these were compared, dimethyl trisulfide (17 ng/µL) and dimethyl disulfide (11 ng/µL) were found to be emitted in higher concentrations. The roles of these compounds and nonanal as core volatiles for cadaver detection or postmortem time determination and their correlation to the stages of decay and the accumulated degree days are discussed.