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Electrospinning is a simple method for obtaining nanofibers. They possess distinctive ‎properties such as large surface area to weight and high porosity, which make them attractive for many ‎applications. In this research, the electrospinning technique was used to obtain non-woven mats of ‎fibers using polylactic acid (PLA) with the addition of curcumin powder and clove oil as natural ‎antibacterial materials. For the preparation of mats, a 10% polylactic acid solution was prepared in ‎a mixture of acetone and dimethylformamide. Natural materials were added to the ‎polymer solution in several concentrations of 1, 3, 5, 7, and 10%. The viscosity of these solutions was ‎measured. The samples were then prepared using a locally manufactured ‎electro-spinner. Using a scanning electron microscope, it was ‎found that the average diameter of the fibers of the polylactic acid sample is 177 nm. It is noted ‎that the diameters of the fibers produced using polymer and natural materials have larger ‎diameters, especially when curcumin is added, as the fiber diameters range between 485 and 764 nm. ‎The activity of the produced samples against two types of Gram-negative and Gram-positive bacteria ‎was tested. It was found that the polymer-only sample did not have any resistance to the bacteria. ‎While samples containing natural materials showed antibacterial activity against both types of bacteria.‎ It was noted that this activity increased with the increase in the concentration of the additive, which ‎produced ‎an increase in the diameter of the inhibition zone, which makes it suitable for medical and ‎other ‎applications.‎

Abstract Local tissue damage following snakebite envenoming remains a poorly researched area. To develop better strategies to treat snakebites, it is critical to understand the mechanisms through which venom toxins induce envenomation effects including local tissue damage. Here, we demonstrate how the venoms of two medically important Indian snakes (Russell's viper and cobra) affect human skeletal muscle using a cultured human myoblast cell line. The data suggest that both venoms affect the viability of myoblasts. Russell’s viper venom reduced the total number of cells, their migration, and the area of focal adhesions. It also suppressed myogenic differentiation and induced muscle atrophy. While cobra venom decreased the viability, it did not largely affect cell migration and focal adhesions. Cobra venom affected the formation of myotubes and induced atrophy. Cobra venom-induced atrophy could not be reversed by small molecule inhibitors such as varespladib (a phospholipase A2 inhibitor) and prinomastat (a metalloprotease inhibitor), and soluble activin type IIb receptor (a molecule used to promote regeneration of skeletal muscle), although the antivenom (raised against the Indian ‘Big Four’ snakes) has attenuated the effects. However, all these molecules rescued the myotubes from Russell’s viper venom-induced atrophy. This study demonstrates key steps in the muscle regeneration process that are affected by both Indian Russell’s viper and cobra venoms and offers insights into the potential causes of clinical features displayed in envenomed victims. Further research is required to investigate the molecular mechanisms of venom-induced myotoxicity under in vivo settings and develop better therapies for snakebite-induced muscle damage.

The study deals with amount of sediment yield in flooding condition in reservoir of Dewerige dam .Since the Dewerige River is seasonal river, therefore the study achievement during dry season. The study aims to calculate amount of siltation which accumulated in reservoir and its impact on reservoir storage capacity, thus reducing economic life time of the dam. Topographic map (dwg file) considered the base map .The final field survey in field work during January 2017 was measured thickness of siltation in reservoir. By chosen cross-sections area in channel of Dewerige River based on Trapezoidal rule the volume of siltation can be calculated. Finally, siltation volume used to calculate: rate of sediment, economic life time of the dam, sediment yield and specific sediment yield in reservoir of Dewerige dam. Deposition of sediments in reservoir assumed as 514000 m3 during designing stage report of Ministry of Irrigation, the general commission of dams and reservoirs, nevertheless the study proved sediment yield is 8242950.6 m3 /y that is a very big amount of sediment for a small dam reservoir. Economic life time take into account during designing stage was 50 years, while the economic life time depend upon this study is 3 years. [ABSTRACT FROM AUTHOR]

Envenomation by the Indian ornamental tarantula (Poecilotheria regalis) is medically relevant to humans, both in its native India and worldwide, where they are kept as pets. Muscle-related symptoms such as cramps and pain are commonly reported in humans following envenomation by this species. There is no specific treatment, including antivenom, for its envenomation. Moreover, the scientific knowledge of the impact of this venom on skeletal muscle function is highly limited. Therefore, we carried out this study to better understand the myotoxic properties of Poecilotheria regalis venom by determining its effects in cultured myoblasts and in the tibialis anterior muscle in mice. While there was no effect found on undifferentiated myoblasts, the venom affected differentiated multinucleated myotubes resulting in the reduction of fusion and atrophy of myotubes. Similarly, intramuscular administration of this venom in the tibialis anterior muscle in mice resulted in extensive muscle damage on day 5. However, by day 10, the regeneration was evident, and the regeneration process continued until day 20. Nevertheless, some tissue abnormalities including reduced dystrophin expression and microthrombi presence were observed on day 20. Overall, this study demonstrates the ability of this venom to induce significant muscle damage and affect its regeneration in the early stages. These data provide novel mechanistic insights into this venom-induced muscle damage and guide future studies to isolate and characterise individual toxic component(s) that induce muscle damage and their significance in developing better therapeutics.

The European Commission have just stated that hydrogen would play a major role in the economic recovery of post-COVID-19 EU countries. Hydrogen is recognised as one of the key players in a fossil fuel-free world in decades to come. However, commercially practiced pathways to hydrogen production todays, are associated with a considerable amount of carbon emissions. The Paris Climate Change Agreement has set out plans for an international commitment to reduce carbon emissions within the forthcoming decades. A sustainable hydrogen future would only be achievable if hydrogen production is “designed” to capture such emissions. Today, nearly 98% of global hydrogen production relies on the utilisation of fossil fuels. Among these, steam methane reforming (SMR) boasts the biggest share of nearly 50% of the global generation. SMR processes correspond to a significant amount of carbon emissions at various points throughout the process. Despite the dark side of the SMR processes, they are projected to play a major role in hydrogen production by the first half of this century. This that a sustainable, yet clean short/medium-term hydrogen production is only possible by devising a plan to efficiently capture this co-produced carbon as stated in the latest International Energy Agency (IEA) reports. Here, we have carried out an in-depth technical review of the processes employed in sorption-enhanced steam methane reforming (SE-SMR), an emerging technology in low-carbon SMR, for combined carbon capture and hydrogen production. This paper aims to provide an in-depth review on two key challenging elements of SE-SMR i.e. the advancements in catalysts/adsorbents preparation, and current approaches in process synthesis and optimisation including the employment of artificial intelligence in SE-SMR processes. To the best of the authors’ knowledge, there is a clear gap in the literature where the above areas have been scrutinised in a systematic and coherent fashion. The gap is even more pronounced in the application of AI in SE-SMR technologies. As a result, this work aims to fill this gap within the scientific literature.

Abstract Background The relationship between the tissue injury healing response and development of heterotopic ossification (HO) is poorly understood. Here we compare a rat blast model and human traumatized muscle from a blast injury to study the early signatures of osteogenesis and fibrosis during the formation of HO. Methods Rat and human tissues were characterized using histology, scanning electron microscopy, immunohistochemistry, as well as gene and protein expression analysis. Additionally, animals and humans were assessed radiographically for HO formation following injury. Results Markers of bone formation were dramatically increased in tissue samples from both humans and rats, and both displayed increased fibroproliferative regions within the injured tissues and elevated expression of markers of tissue fibrosis such as TGF-β1, Fibronectin, SMAD3 and PAI-1. Markers of inflammation and fibrosis (ACTA, TNFα, BMP1 and BMP3) were elevated at the RNA level in both rat and human samples. By day 42, bone formation in the rat blast model appeared similar in radiographs compared to human patients who progressed to develop post-traumatic HO. Conclusions Our data demonstrates that a similar early fibrotic response is evident in both the rat blast model and the human tissues following a traumatic injury and demonstrates the relevance of this animal model for future translational studies.

Homogeneity is a concept used by grammarians to explain several grammatical provisions dispersed amongst multiple fields. This research paper examines this concept and its impact on Arabic grammar at the syntactical and interpretation levels and how it relates to other rules. This study compiles locations and texts in which the term is mentioned and others used by grammarians to relay the same concept. The impact of this issue is clear in its ability to display the pivotal impact of the concept of homogeneity in the Arabic linguistic rhetoric. Despite that fact that it appears clearly in the field of comparative literature, there is still some room for further investigation in the grammatical discourse due to lack of attention granted to it. This research paper includes an introduction and three research points, the first is for the term (Almoshakalah). The second which investigates the theoretical framework of homogeneity and the third which investigates various forms of grammatical and linguistical homogeneity after which the conclusion states essential research points.

This research focuses on studying the effect of electrospinning device room temperature on PVA nano-fibers diameter made by electrospinning device depositing on rotating drum. In this research, the preparation of nano-non woven fabrics by electro-spun polymer solution of poly vinyl alcohol using distilled water as solvent in temperature range [15-35]C° was achieved. The relative humidity 40%, polymer solution concentration 14% wt., voltage 22KV, tip-to-collector distance 10 Cm. The results of the five scanned samples by scanning electron microscope (SEM) indicated that: by increasing room temperature the diameter of nano-fibers decreased. Also it is important to mention that the range of room temperature shouldn't be exceeded in order not to close the orifice of the needle because of the evaporation of the droplet solvent changing into solid polymer.

Background: The prevalence of new diseases and increased number of patient referring to health care centers cause more medical malpractice. This study aimed to identify the frequency and reasons of medical errors in cases referred to Isfahan Legal Medicine Commission during 2005-2009. It also tried to provide ways to reduce such errors. Methods: In a descriptive, cross-sectional study, 380 cases of medical errors referred to Isfahan Legal Medicine Commission from 2005-2009 were evaluated. Due to inaccessibility and incompleteness of information, and also uselessness of some cases, 352 cases were finally investigated. The data was collected by a checklist whose validity was obtained by the opinions of experts in several stages. Data collection methods were resource review and observation. Data was analyzed by SPSS11. Results: The 352 studied records included claims from individuals. Among physicians and surgeons, general practitioners were claimed more than others (15.3%). Anesthesiologists, obstetrics and gynecologists, general surgeons and orthopedic specialists were claimed in 9.3%, 8.3%, 7.6%, and 7.4% of cases, respectively. Among other health care professionals, nurses had the highest frequency of claims (9.8%). They were in the second rank of total sentences. In addition, 36.9% of sentences led to conviction among which general practitioners were in the first place (15.3%). The average age of staff was 43 years. The highest frequency of claims (23.2%) was observed in the age group of 35-40 years. Males constituted 68.2% of the medical staff. Among the 23 cases of administrative staff errors, 91.3% cases led to conviction. In 35.2% of cases, negligence was the main reason of medical malpractice. In 46.4% of the issued convictions, a blood money of 1-10% was determined. Conclusion: Our results showed that patients' claims of medical staff were increased during the past 5 years. In fact, while 62 cases belonged to 2006, 108 cases were reported in 2010. Although medical errors are inconsiderable when the delivered health care services are concerned, identifying their reasons and a proper health care management would lead to higher quality of provided services. Key Words: Malpractice; Forensic Medicine; Health Services.

Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder characterized by challenges in social communication as well as repetitive or restrictive behaviors. Many genetic associations with ASD have been identified, but most associations occur in a fraction of the ASD population. Here, we searched for eQTL-associated DNA variants with significantly different allele distributions between ASD-affected and control. Thirty significant DNA variants associated with 174 tissue-specific eQTLs from ASD individuals in the SPARK project were identified. Several significant variants fell within brain-specific regulatory regions or had been associated with a significant change in gene expression in the brain. These eQTLs are a new class of biomarkers that could control the myriad of brain and non-brain phenotypic traits seen in ASD-affected individuals., (© 2022. The Author(s).)

Schwannoma is a benign, encapsulated, slow-growing and generally solitary tumour that arise from Schwann cells of the peripheral nerve sheath. Approximately 1-12% occur intraorally with the tongue being one of the site. We report a case of schwannoma of the tongue in a 33-year-old male patient. It presented as a slow growing mass of the tongue. We did the surgical excision of the tumour with the help of CO 2 laser, and the patient is in follow-up and is asymptomatic from the last 7 months., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© Association of Otolaryngologists of India 2021.)

Spindle Cell Carcinoma (SpCC) is a rare neoplasm that occurs mainly in the upper aero-digestive tract, mostly in larynx. We present a case of SpCC arising from nasal septum. Surgical excision was done with postoperative chemo-radiotherapy and the patient is in follow-up and is asymptomatic from the last 24 months., Competing Interests: Conflicts of interestThe authors have no conflicts of interest to disclose., (© Association of Otolaryngologists of India 2021.)

Receptor-ligand interactions on the plasma membrane regulate cellular communication and play a key role in viral infection. Despite representing main targets for drug development, the characterization of these interactions remains challenging in part due to the dearth of optimal technologies. Here, we build a comprehensive library of human proteins engineered for controlled cell surface expression. Coupled to tetramer-based screening for increased binding avidity, we develop a high throughput cell-based platform that enables systematic interrogation of receptor-ligand interactomes. Using this technology, we characterize the cell surface proteins targeted by the receptor binding domain (RBD) of the SARS-CoV spike protein. Host factors that specifically bind to SARS CoV-2 but not SARS CoV RBD are identified, including proteins that are expressed in the nervous system or olfactory epithelium. Remarkably, our results show that Contactin-1, a previously unknown SARS CoV-2 spike-specific receptor that is upregulated in COVID-19 patients, significantly enhances ACE2-dependent pseudotyped virus infection. Starting from a versatile platform to characterize cell surface interactomes, this study uncovers host factors specifically targeted by SARS CoV-2, information that may help design improved therapeutic strategies against COVID-19., (© 2022. The Author(s).)

New therapeutics and combination regimens have led to marked clinical improvements for the treatment of a subset of colorectal cancer. Immune checkpoint inhibitors have shown clinical efficacy in patients with mismatch-repair-deficient or microsatellite instability-high (MSI-H) metastatic colorectal cancer (mCRC). However, patients with microsatellite-stable (MSS) or low levels of microsatellite instable (MSI-L) colorectal cancer have not benefited from these immune modulators, and the survival outcome remains poor for the majority of patients diagnosed with mCRC. In this article, we describe the discovery of a novel T-cell-dependent bispecific antibody (TDB) targeting tumor-associated antigen LY6G6D, LY6G6D-TDB, for the treatment of colorectal cancer. RNAseq analysis showed that LY6G6D was differentially expressed in colorectal cancer with high prevalence in MSS and MSI-L subsets, whereas LY6G6D expression in normal tissues was limited. IHC confirmed the elevated expression of LY6G6D in primary and metastatic colorectal tumors, whereas minimal or no expression was observed in most normal tissue samples. The optimized LY6G6D-TDB, which targets a membrane-proximal epitope of LY6G6D and binds to CD3 with high affinity, exhibits potent antitumor activity both in vitro and in vivo. In vitro functional assays show that LY6G6D-TDB-mediated T-cell activation and cytotoxicity are conditional and target dependent. In mouse xenograft tumor models, LY6G6D-TDB demonstrates antitumor efficacy as a single agent against established colorectal tumors, and enhanced efficacy can be achieved when LY6G6D-TDB is combined with PD-1 blockade. Our studies provide evidence for the therapeutic potential of LY6G6D-TDB as an effective treatment option for patients with colorectal cancer., (©2022 The Authors; Published by the American Association for Cancer Research.)

The mechanisms that coordinate cellular gene expression are highly complex and intricately interconnected. Thus, it is necessary to move beyond a fully reductionist approach to understanding genetic information flow and begin focusing on the networked connections between genes that organize cellular function. Continued advancements in computational hardware, coupled with the development of gene correlation network algorithms, provide the capacity to study networked interactions between genes rather than their isolated functions. For example, gene coexpression networks are used to construct gene relationship networks using linear metrics such as Spearman or Pearson correlation. Recently, there have been tools designed to deepen these analyses by differentiating between intrinsic vs extrinsic noise within gene expression values, identifying different modules based on tissue phenotype, and capturing potential nonlinear relationships. In this report, we introduce an algorithm with a novel application of image-based segmentation modalities utilizing blob detection techniques applied for detecting bigenic edges in a gene expression matrix. We applied this algorithm called EdgeCrafting to a bulk RNA-sequencing gene expression matrix comprised of a healthy kidney and cancerous kidney data. We then compared EdgeCrafting against 4 other RNA expression analysis techniques: Weighted Gene Correlation Network Analysis, Knowledge Independent Network Construction, NetExtractor, and Differential gene expression analysis., (© The Author(s) 2022. Published by Oxford University Press on behalf of Genetics Society of America.)

The cancer-associated fibroblast (CAF) marker podoplanin (PDPN) is generally correlated with poor clinical outcomes in cancer patients and thus represents a promising therapeutic target. Despite its biomedical relevance, basic aspects of PDPN biology such as its cellular functions and cell surface ligands remain poorly uncharacterized, thus challenging drug development. Here, we utilize a high throughput platform to elucidate the PDPN cell surface interactome, and uncover the neutrophil protein CD177 as a new binding partner. Quantitative proteomics analysis of the CAF phosphoproteome reveals a role for PDPN in cell signaling, growth and actomyosin contractility, among other processes. Moreover, cellular assays demonstrate that CD177 is a functional antagonist, recapitulating the phenotype observed in PDPN-deficient CAFs. In sum, starting from the unbiased elucidation of the PDPN co-receptome, our work provides insights into PDPN functions and reveals the PDPN/CD177 axis as a possible modulator of fibroblast physiology in the tumor microenvironment., Competing Interests: Y.S., S.G., V.C.P., C.C., J.R.L., and S.J.T. are Genentech employees and own shares in the Genentech/Roche group. J.L.A, S.K., B.H., E.V., S.M.P., A.A.P., L.G. and N.M.M. were employees of Roche when the data in this paper was generated.

Myeloid cells encounter stromal cells and their matrix determinants on a continual basis during their residence in any given organ. Here, we examined the impact of the collagen receptor LAIR1 on myeloid cell homeostasis and function. LAIR1 was highly expressed in the myeloid lineage and enriched in non-classical monocytes. Proteomic definition of the LAIR1 interactome identified stromal factor Colec12 as a high-affinity LAIR1 ligand. Proteomic profiling of LAIR1 signaling triggered by Collagen1 and Colec12 highlighted pathways associated with survival, proliferation, and differentiation. Lair1 -/- mice had reduced frequencies of Ly6C - monocytes, which were associated with altered proliferation and apoptosis of non-classical monocytes from bone marrow and altered heterogeneity of interstitial macrophages in lung. Myeloid-specific LAIR1 deficiency promoted metastatic growth in a melanoma model and LAIR1 expression associated with improved clinical outcomes in human metastatic melanoma. Thus, monocytes and macrophages rely on LAIR1 sensing of stromal determinants for fitness and function, with relevance in homeostasis and disease., Competing Interests: Declarations of interests All authors are stockholders of Genentech/Roche except B.H., A.W., E.V., and S.C., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Purpose: Immune checkpoint inhibitors (ICI) are highly effective in several cancer entities, but also invoke a variety of immune-related adverse events (irAE). These are mostly reversible, but can be life-threatening or even fatal. Currently, the pathogenesis is not fully understood, but crucial for effective treatment. Prediction and early detection of irAE could be facilitated and treatment optimized if relevant biomarkers and effector mechanisms were better characterized., Methods: This study included a total of 45 irAE in patients with metastatic melanoma who were treated with ICI. All patients underwent a complete work-up with exclusion of other causes. Longitudinal blood samples were analyzed for a panel of soluble markers and compared to baseline and to patients who did not experience any irAE. Measurements included LDH, interleukin (IL)-6, IL-1β, IL-17, C-reactive protein (CRP) and tumor necrosis factor (TNF)-alpha as well as tumor markers S100 and melanoma inhibitory activity (MIA)., Results: During the early onset of irAE increases in serum IL-6 (from mean 24.4 pg/ml at baseline to 51.0 pg/ml; p = 0.003) and CRP (from mean 7.0 mg/l at baseline to 17.7 mg/l; p = 0.001) and a decrease in MIA (from mean 5.4 pg/ml at baseline to 4.8 pg/ml; p = 0.035) were detected. No changes in IL-17 were noted. These effects were observed for irAE of different organ systems., Conclusion: Increases of a combination of IL-6 and CRP serum levels can be used for the early detection of irAE and tailored management. Interestingly, changes in MIA serum levels also correlate with irAE onset.

Protein kinase R (PKR) is a key pattern recognition receptor of the innate immune pathway. PKR is activated by double-stranded RNA (dsRNA) that is often produced during viral genome replication and transcription. PKR contains two tandem double-stranded RNA binding domains at the N-terminus, dsRBD1 and dsRBD2, and a C-terminal kinase domain. In the canonical model for activation, RNAs that bind multiple PKRs induce dimerization of the kinase domain that promotes an active conformation. However, there is evidence that dimerization of the kinase domain is not sufficient to mediate activation and PKR activation is modulated by the RNA-binding mode. dsRBD2 lacks most of the consensus RNA-binding residues, and it has been suggested to function as a modulator of PKR activation. Here, we demonstrate that dsRBD2 regulates PKR activation and identify the N-terminal helix as a critical region for modulating kinase activity. Mutations in dsRBD2 that have minor effects on overall dsRNA-binding affinity strongly inhibit the activation of PKR by dsRNA. These mutations also inhibit RNA-independent PKR activation. These data support a model where dsRBD2 has evolved to function as a regulator of the kinase., Competing Interests: The authors declare no competing financial interest., (© 2021 The Authors. Published by American Chemical Society.)

The human brain is a complex organ that consists of several regions each with a unique gene expression pattern. Our intent in this study was to construct a gene co-expression network (GCN) for the normal brain using RNA expression profiles from the Genotype-Tissue Expression (GTEx) project. The brain GCN contains gene correlation relationships that are broadly present in the brain or specific to thirteen brain regions, which we later combined into six overarching brain mini-GCNs based on the brain's structure. Using the expression profiles of brain region-specific GCN edges, we determined how well the brain region samples could be discriminated from each other, visually with t-SNE plots or quantitatively with the Gene Oracle deep learning classifier. Next, we tested these gene sets on their relevance to human tumors of brain and non-brain origin. Interestingly, we found that genes in the six brain mini-GCNs showed markedly higher mutation rates in tumors relative to matched sets of random genes. Further, we found that cortex genes subdivided Head and Neck Squamous Cell Carcinoma (HNSC) tumors and Pheochromocytoma and Paraganglioma (PCPG) tumors into distinct groups. The brain GCN and mini-GCNs are useful resources for the classification of brain regions and identification of biomarker genes for brain related phenotypes.

Bigenic expression relationships are conventionally defined based on metrics such as Pearson or Spearman correlation that cannot typically detect latent, non-linear dependencies or require the relationship to be monotonic. Further, the combination of intrinsic and extrinsic noise as well as embedded relationships between sample sub-populations reduces the probability of extracting biologically relevant edges during the construction of gene co-expression networks (GCNs). In this report, we address these problems via our NetExtractor algorithm. NetExtractor examines all pairwise gene expression profiles first with Gaussian mixture models (GMMs) to identify sample sub-populations followed by mutual information (MI) analysis that is capable of detecting non-linear differential bigenic expression relationships. We applied NetExtractor to brain tissue RNA profiles from the Genotype-Tissue Expression (GTEx) project to obtain a brain tissue specific gene expression relationship network centered on cerebellar and cerebellar hemisphere enriched edges. We leveraged the PsychENCODE pre-frontal cortex (PFC) gene regulatory network (GRN) to construct a cerebellar cortex (cerebellar) GRN associated with transcriptionally active regions in cerebellar tissue. Thus, we demonstrate the utility of our NetExtractor approach to detect biologically relevant and novel non-linear binary gene relationships., (Copyright © 2020 Husain et al.)

We introduce the Transcriptome State Perturbation Generator (TSPG) as a novel deep-learning method to identify changes in genomic expression that occur between tissue states using generative adversarial networks. TSPG learns the transcriptome perturbations from RNA-sequencing data required to shift from a source to a target class. We apply TSPG as an effective method of detecting biologically relevant alternate expression patterns between normal and tumor human tissue samples. We demonstrate that the application of TSPG to expression data obtained from a biopsy sample of a patient's kidney cancer can identify patient-specific differentially expressed genes between their individual tumor sample and a target class of healthy kidney gene expression. By utilizing TSPG in a precision medicine application in which the patient sample is not replicated (i.e., n = 1 ), we present a novel technique of determining significant transcriptional aberrations that can be used to help identify potential targeted therapies., Competing Interests: The authors declare no competing interests., (© 2020 The Authors.)

Cell surface receptors and their interactions play a central role in physiological and pathological signaling. Despite its clinical relevance, the immunoglobulin superfamily (IgSF) remains uncharacterized and underrepresented in databases. Here, we present a systematic extracellular protein map, the IgSF interactome. Using a high-throughput technology to interrogate most single transmembrane receptors for binding to 445 IgSF proteins, we identify over 500 interactions, 82% previously undocumented, and confirm more than 60 receptor-ligand pairs using orthogonal assays. Our study reveals a map of cell-type-specific interactions and the landscape of dysregulated receptor-ligand crosstalk in cancer, including selective loss of function for tumor-associated mutations. Furthermore, investigation of the IgSF interactome in a large cohort of cancer patients identifies interacting protein signatures associated with clinical outcome. The IgSF interactome represents an important resource to fuel biological discoveries and a framework for understanding the functional organization of the surfaceome during homeostasis and disease, ultimately informing therapeutic development., Competing Interests: Declaration of Interests B.H., Y. Senbabaoglu, I.L., T.A.A., B.W., C.B., Y.F., P.C., A.W.W., L.C.G., S.M., S.J.T., J.R.L., and N.M.-M. are Genentech employees and own shares in the Roche group., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Receptors expressed on the plasma membrane and their interacting partners critically regulate cellular communication during homeostasis and disease, and as such represent main therapeutic targets. Despite its importance for drug development, receptor-ligand proteomics has remained a daunting field, in part because of the challenges associated to the study of membrane-expressed proteins. Here, to enable sensitive detection of receptor-ligand interactions in high throughput, we implement a new platform, the Conditioned Media AlphaScreen, for interrogation of a library consisting of most single transmembrane human proteins. Using this method to study key immune receptors, we identify and further validate the interleukin receptor IL20RA as the first binding partner for the checkpoint inhibitor B7-H3. Further, KIR2DL5, a natural killer cell protein that had remained orphan, is uncovered as a functional binding partner for the poliovirus receptor (PVR). This interaction is characterized using orthogonal assays, which demonstrate that PVR specifically engages KIR2DL5 on natural killer cells leading to inhibition of cytotoxicity. Altogether, these results reveal unappreciated links between protein families that may importantly influence receptor-driven functions during disease. Applicable to any target of interest, this technology represents a versatile and powerful approach for elucidation of receptor-ligand interactomes, which is essential to understand basic aspects of the biology of the plasma membrane proteins and ultimately inform the development of novel therapeutic strategies., (© 2019 Husain et al.)

Gene co-expression networks (GCNs) are constructed from Gene Expression Matrices (GEMs) in a bottom up approach where all gene pairs are tested for correlation within the context of the input sample set. This approach is computationally intensive for many current GEMs and may not be scalable to millions of samples. Further, traditional GCNs do not detect non-linear relationships missed by correlation tests and do not place genetic relationships in a gene expression intensity context. In this report, we propose EdgeScaping, which constructs and analyzes the pairwise gene intensity network in a holistic, top down approach where no edges are filtered. EdgeScaping uses a novel technique to convert traditional pairwise gene expression data to an image based format. This conversion not only performs feature compression, making our algorithm highly scalable, but it also allows for exploring non-linear relationships between genes by leveraging deep learning image analysis algorithms. Using the learned embedded feature space we implement a fast, efficient algorithm to cluster the entire space of gene expression relationships while retaining gene expression intensity. Since EdgeScaping does not eliminate conventionally noisy edges, it extends the identification of co-expression relationships beyond classically correlated edges to facilitate the discovery of novel or unusual expression patterns within the network. We applied EdgeScaping to a human tumor GEM to identify sets of genes that exhibit conventional and non-conventional interdependent non-linear behavior associated with brain specific tumor sub-types that would be eliminated in conventional bottom-up construction of GCNs. Edgescaping source code is available at https://github.com/bhusain/EdgeScaping under the MIT license., Competing Interests: The authors have declared that no competing interests exist.

Secreted factors, membrane-tethered receptors, and their interacting partners are main regulators of cellular communication and initiation of signaling cascades during homeostasis and disease, and as such represent prime therapeutic targets. Despite their relevance, these interaction networks remain significantly underrepresented in current databases; therefore, most extracellular proteins have no documented binding partner. This discrepancy is primarily due to the challenges associated with the study of the extracellular proteins, including expression of functional proteins, and the weak, low affinity, protein interactions often established between cell surface receptors. The purpose of this method is to describe the printing of a library of extracellular proteins in a microarray format for screening of protein-protein interactions. To enable detection of weak interactions, a method based on multimerization of the query protein under study is described. Coupled to this microbead-based multimerization approach for increased multivalency, the protein microarray allows robust detection of transient protein-protein interactions in high throughput. This method offers a rapid and low sample consuming-approach for identification of new interactions applicable to any extracellular protein. Protein microarray printing and screening protocol are described. This technology will be useful for investigators seeking a robust method for discovery of protein interactions in the extracellular space.

Bispecific antibodies have moved from being an academic curiosity with therapeutic promise to reality, with two molecules being currently commercialized (Hemlibra ® and Blincyto ® ) and many more in clinical trials. The success of bispecific antibodies is mainly due to the continuously growing number of mechanisms of actions (MOA) they enable that are not accessible to monoclonal antibodies. One of the earliest MOA of bispecific antibodies and currently the one with the largest number of clinical trials is the redirecting of the cytotoxic activity of T-cells for oncology applications, now extending its use in infective diseases. The use of bispecific antibodies for crossing the blood-brain barrier is another important application because of its potential to advance the therapeutic options for neurological diseases. Another noteworthy application due to its growing trend is enabling a more tissue-specific delivery or activity of antibodies. The different molecular solutions to the initial hurdles that limited the development of bispecific antibodies have led to the current diverse set of bispecific or multispecific antibody formats that can be grouped into three main categories: IgG-like formats, antibody fragment-based formats, or appended IgG formats. The expanded applications of bispecific antibodies come at the price of additional challenges for clinical development. The rising complexity in their structure may increase the risk of immunogenicity and the multiple antigen specificity complicates the selection of relevant species for safety assessment.

RNA-activated protein kinase (PKR) is a key component of the interferon-induced antiviral pathway in higher eukaryotes. Upon recognition of viral dsRNA, PKR is activated via dimerization and autophosphorylation. PKR contains two N-terminal dsRNA binding domains (dsRBD) and a C-terminal kinase domain. The dsRBDs and the kinase are separated by a long, unstructured ∼80-amino acid linker in the human enzyme. The length of the N-terminal portion of the linker varies among PKR sequences, and it is completely absent in one ortholog. Here, we characterize the effects of deleting the variable region from the human enzyme to produce PKRΔV. The linker deletion results in quantitative but not qualitative changes in catalytic activity, RNA binding, and conformation. PKRΔV is somewhat more active and exhibits more cooperative RNA binding. As we previously observed for the full-length enzyme, PKRΔV is flexible in solution and adopts a range of compact and extended conformations. The conformational ensemble is biased toward compact states that might be related to weak interactions between the dsRBD and kinase domains. PKR retains RNA-induced autophosphorylation upon complete removal of the linker, indicating that the C-terminal, basic region is also not required for activity.

PKR is a member of the eIF2α family of protein kinases that inhibit translational initiation in response to stress stimuli and functions as a key mediator of the interferon-induced antiviral response. PKR contains a dsRNA binding domain that binds to duplex regions present in viral RNAs, resulting in kinase activation and autophosphorylation. An emerging theme in the regulation of protein kinases is the allosteric linkage of dimerization and activation. The PKR kinase domain forms a back-to-back parallel dimer that is implicated in activation. We have developed a sensitive homo-Förster resonance energy transfer assay for kinase domain dimerization to directly probe the relationship among RNA binding, activation, and dimerization. In the case of perfect duplex RNAs, dimerization is correlated with activation and dsRNAs containing 30 bp or more efficiently induce kinase domain dimerization and activation. However, more complex duplex RNAs containing a 10-15 bp 2'-O-methyl RNA barrier produce kinase dimers but do not activate. Similarly, inactivating mutations within the PKR dimer interface that disrupt key electrostatic and hydrogen binding interactions fail to abolish dimerization. Our data support a model in which activating RNAs induce formation of a back-to-back parallel PKR kinase dimer whereas nonactivating RNAs either fail to induce dimerization or produce an alternative, inactive dimer configuration, providing an additional mechanism for distinguishing between host and pathogen RNA.