scholarly journals ATRT-24. CELL SURFACE PROTEOME ANALYSIS OF ATRT IDENTIFIES TARGETS FOR IMMUNOTHERAPY

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii280-iii280
Author(s):  
Allison Cole ◽  
Eric Hoffmeyer ◽  
Marco Zanini ◽  
Rajeev Vibhakar ◽  
Anandani Nellan ◽  
...  
Keyword(s):  
Large Scale ◽  
Proteome Analysis ◽  
Surface Expression ◽  
Surface Proteins ◽  
Dna And Rna ◽  
Single Deletion ◽  
Surface Receptor ◽  
Atypical Teratoid ◽  
And Function ◽  
Surface Proteome

Abstract Atypical teratoid rhabdoid tumor (ATRT) is a rare and fast-growing childhood tumor of the brain and spinal cord. While the recent advances in DNA and RNA sequencing have given deep insights into the biology of cancer, about 90% of ATRTs harbor a single deletion which leads to uncontrolled tumor growth. The lack of targetable genetic abnormalities in ATRT makes it a tough target for therapy and hence radical new approaches are required to develop a treatment. In many cases, the gene expression profile alone DOES NOT represent the presence of the gene product on the surface and cannot detect post-translational modifications such as the addition of sugars which are essential for the interaction of surface proteins with the tumor microenvironment. The ability to escape from surveillance by the immune system is regarded as one of the essential hallmarks of cancer cells. Here we carried out a comprehensive unbiased large-scale surface receptor profiling using high throughput multicolor flow cytometry on surgically resected ATRT patient samples, primary ATRT cell lines, and patient-derived xenograft models. By multiplexing primary samples with antibodies for CD31, CD45, CD11b, CCR2, Cx3cr1, and CD4, and CD8 we eliminated endothelial and immune cells from analysis while also identifying immune populations within the tumor. We identified increased surface expression of CD44, CD146, CD59, CD151, and CD276. These were validated in our screening of primary tumor samples. A combination of CAR-T cell and function-blocking monoclonal antibody approaches have been tested to verify the proof of principle of this approach.

scholarly journals ASTN2 modulates synaptic strength by trafficking and degradation of surface proteins

2018 ◽  
Author(s):  
Hourinaz Behesti ◽  
Taylor Fore ◽  
Peter Wu ◽  
Zachi Horn ◽  
Mary Leppert ◽  
...  
Keyword(s):  
Surface Protein ◽  
Surface Expression ◽  
Autism Spectrum ◽  
Copy Number Variations ◽  
Surface Proteins ◽  
Synaptic Activity ◽  
Synaptic Proteins ◽  
Dynamic Regulation ◽  
Cerebellar Purkinje Cells ◽  
And Function

AbstractSurface protein dynamics dictate synaptic connectivity and function in neuronal circuits. ASTN2, a gene disrupted by copy number variations (CNVs) in neurodevelopmental disorders, including autism spectrum, was previously shown to regulate the surface expression of ASTN1 in glial-guided neuronal migration. Here, we demonstrate that ASTN2 binds to and regulates the surface expression of multiple synaptic proteins in post-migratory neurons by endocytosis, resulting in modulation of synaptic activity. In cerebellar Purkinje cells (PCs), by immuno-gold electron microscopy, ASTN2 localizes primarily to endocytic and autophagocytic vesicles in the cell soma and in subsets of dendritic spines. Overexpression of ASTN2 in PCs, but not of ASTN2 lacking the FNIII-domain commonly disrupted by CNVs in patients including in a family presented here, increases inhibitory and excitatory postsynaptic activity and reduces levels of ASTN2 binding partners. Our data suggest a fundamental role for ASTN2 in dynamic regulation of surface proteins by endocytic trafficking and protein degradation.

scholarly journals Molecular pathogenesis of human CD59 deficiency

2018 ◽  
Vol 4 (6) ◽  
pp. e280 ◽  
Author(s):  
Netanel Karbian ◽  
Yael Eshed-Eisenbach ◽  
Adi Tabib ◽  
Hila Hoizman ◽  
B. Paul Morgan ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Cell Surface ◽  
Chronic Inflammatory Demyelinating Polyneuropathy ◽  
Membrane Attack Complex ◽  
Surface Expression ◽  
Surface Proteins ◽  
Cell Surface Expression ◽  
Congenital Deficiency ◽  
Recurrent Strokes ◽  
And Function

ObjectiveTo characterize all 4 mutations described for CD59 congenital deficiency.MethodsThe 4 mutations, p.Cys64Tyr, p.Asp24Val, p.Asp24Valfs*, and p.Ala16Alafs*, were described in 13 individuals with CD59 malfunction. All 13 presented with recurrent Guillain-Barré syndrome or chronic inflammatory demyelinating polyneuropathy, recurrent strokes, and chronic hemolysis. Here, we track the molecular consequences of the 4 mutations and their effects on CD59 expression, localization, glycosylation, degradation, secretion, and function. Mutants were cloned and inserted into plasmids to analyze their expression, localization, and functionality.ResultsImmunolabeling of myc-tagged wild-type (WT) and mutant CD59 proteins revealed cell surface expression of p.Cys64Tyr and p.Asp24Val detected with the myc antibody, but no labeling by anti-CD59 antibodies. In contrast, frameshift mutants p.Asp24Valfs* and p.Ala16Alafs* were detected only intracellularly and did not reach the cell surface. Western blot analysis showed normal glycosylation but mutant-specific secretion patterns. All mutants significantly increased MAC-dependent cell lysis compared with WT. In contrast to CD59 knockout mice previously used to characterize phenotypic effects of CD59 perturbation, all 4 hCD59 mutations generate CD59 proteins that are expressed and may function intracellularly (4) or on the cell membrane (2). None of the 4 CD59 mutants are detected by known anti-CD59 antibodies, including the 2 variants present on the cell membrane. None of the 4 inhibits membrane attack complex (MAC) formation.ConclusionsAll 4 mutants generate nonfunctional CD59, 2 are expressed as cell surface proteins that may function in non–MAC-related interactions and 2 are expressed only intracellularly. Distinct secretion of soluble CD59 may have also a role in disease pathogenesis.

Scanning tunneling microscopy (STM) of DNA and RNA

1989 ◽  
Vol 47 ◽  
pp. 34-35
Author(s):  
Patricia G. Arscott ◽  
Gil Lee ◽  
Victor A. Bloomfield ◽  
D. Fennell Evans
Keyword(s):  
Molecular Structures ◽  
Inorganic Materials ◽  
Resolving Power ◽  
Scanning Tunneling ◽  
Tunneling Microscopy ◽  
Form And Function ◽  
Dna And Rna ◽  
Calf Thymus ◽  
And Function ◽  
The Relationship

STM is one of the most promising techniques available for visualizing the fine details of biomolecular structure. It has been used to map the surface topography of inorganic materials in atomic dimensions, and thus has the resolving power not only to determine the conformation of small molecules but to distinguish site-specific features within a molecule. That level of detail is of critical importance in understanding the relationship between form and function in biological systems. The size, shape, and accessibility of molecular structures can be determined much more accurately by STM than by electron microscopy since no staining, shadowing or labeling with heavy metals is required, and there is no exposure to damaging radiation by electrons. Crystallography and most other physical techniques do not give information about individual molecules.We have obtained striking images of DNA and RNA, using calf thymus DNA and two synthetic polynucleotides, poly(dG-me5dC)·poly(dG-me5dC) and poly(rA)·poly(rU).

Plant hormones, plant growth regulators

2014 ◽  
Vol 155 (26) ◽  
pp. 1011-1018 ◽  
Author(s):  
György Végvári ◽  
Edina Vidéki
Keyword(s):  
Nervous System ◽  
Growth And Development ◽  
Large Scale ◽  
Essential Role ◽  
Higher Plants ◽  
Structure And Function ◽  
Wide Sense ◽  
Large Scale Integration ◽  
Scale Integration ◽  
And Function

Plants seem to be rather defenceless, they are unable to do motion, have no nervous system or immune system unlike animals. Besides this, plants do have hormones, though these substances are produced not in glands. In view of their complexity they lagged behind animals, however, plant organisms show large scale integration in their structure and function. In higher plants, such as in animals, the intercellular communication is fulfilled through chemical messengers. These specific compounds in plants are called phytohormones, or in a wide sense, bioregulators. Even a small quantity of these endogenous organic compounds are able to regulate the operation, growth and development of higher plants, and keep the connection between cells, tissues and synergy beween organs. Since they do not have nervous and immume systems, phytohormones play essential role in plants’ life. Orv. Hetil., 2014, 155(26), 1011–1018.

scholarly journals Buffalo sperm surface proteome profiling reveals an intricate relationship between innate immunity and reproduction

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Vipul Batra ◽  
Vanya Bhushan ◽  
Syed Azmal Ali ◽  
Parul Sarwalia ◽  
Ankit Pal ◽  
...  
Keyword(s):  
Male Fertility ◽  
Female Reproductive Tract ◽  
Surface Proteins ◽  
Glycosylation Pattern ◽  
Sperm Surface ◽  
Beta Defensins ◽  
Surface Proteome ◽  
Related Proteins ◽  
Sperm Surface Proteins

Abstract Background Low conception rate (CR) despite insemination with morphologically normal spermatozoa is a common reproductive restraint that limits buffalo productivity. This accounts for a significant loss to the farmers and the dairy industry, especially in agriculture-based economies. The immune-related proteins on the sperm surface are known to regulate fertility by assisting the spermatozoa in their survival and performance in the female reproductive tract (FRT). Regardless of their importance, very few studies have specifically catalogued the buffalo sperm surface proteome. The study was designed to determine the identity of sperm surface proteins and to ascertain if the epididymal expressed beta-defensins (BDs), implicated in male fertility, are translated and applied onto buffalo sperm surface along with other immune-related proteins. Results The raw mass spectra data searched against an in-house generated proteome database from UniProt using Comet search engine identified more than 300 proteins on the ejaculated buffalo sperm surface which were bound either by non-covalent (ionic) interactions or by a glycosylphosphatidylinositol (GPI) anchor. The singular enrichment analysis (SEA) revealed that most of these proteins were extracellular with varied binding activities and were involved in either immune or reproductive processes. Flow cytometry using six FITC-labelled lectins confirmed the prediction of glycosylation of these proteins. Several beta-defensins (BDs), the anti-microbial peptides including the BuBD-129 and 126 were also identified amongst other buffalo sperm surface proteins. The presence of these proteins was subsequently confirmed by RT-qPCR, immunofluorescence and in vitro fertilization (IVF) experiments. Conclusions The surface of the buffalo spermatozoa is heavily glycosylated because of the epididymal secreted (glyco) proteins like BDs and the GPI-anchored proteins (GPI-APs). The glycosylation pattern of buffalo sperm-surface, however, could be perturbed in the presence of elevated salt concentration or incubation with PI-PLC. The identification of numerous BDs on the sperm surface strengthens our hypothesis that the buffalo BDs (BuBDs) assist the spermatozoa either in their survival or in performance in the FRT. Our results suggest that BuBD-129 is a sperm-surface BD that could have a role in buffalo sperm function. Further studies elucidating its exact physiological function are required to better understand its role in the regulation of male fertility.

Functional design of glycan-conjugated molecules using a chemoenzymatic approach

2021 ◽  
Author(s):  
Makoto Ogata
Keyword(s):  
Molecular Weight ◽  
Large Scale ◽  
Biological Activities ◽  
Natural Compounds ◽  
Low Molecular Weight ◽  
Functional Design ◽  
Enzymatic Method ◽  
Conjugated Molecules ◽  
Design Synthesis ◽  
And Function

Abstract Carbohydrates play important and diverse roles in the fundamental processes of life. We have established a method for accurately and a large scale synthesis of functional carbohydrates with diverse properties using a unique enzymatic method. Furthermore, various artificial glycan-conjugated molecules have been developed by adding these synthetic carbohydrates to macromolecules and to middle and low molecular weight molecules with different properties. These glycan-conjugated molecules have biological activities comparable to or higher than those of natural compounds, and present unique functions. In this review, several synthetic glycan-conjugated molecules are taken as examples to show design, synthesis and function.

scholarly journals Extracellular Proteome Analysis Shows the Abundance of Histidine Kinase Sensor Protein, DNA Helicase, Putative Lipoprotein Containing Peptidase M75 Domain and Peptidase C39 Domain Protein in Leptospira interrogans Grown in EMJH Medium

Pathogens ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 852
Author(s):  
Abhijit Sarma ◽  
Dhandapani Gunasekaran ◽  
Devasahayam Arokia Balaya Rex ◽  
Thoduvayil Sikha ◽  
Homen Phukan ◽  
...  
Keyword(s):  
Culture Medium ◽  
Proteome Analysis ◽  
Dna Helicase ◽  
Extracellular Proteins ◽  
Surface Proteins ◽  
Secretory Proteins ◽  
Extracellular Proteome ◽  
Cell Pellet ◽  
The Difference ◽  
Leptospira Species

Leptospirosis is a re-emerging form of zoonosis that is caused by the spirochete pathogen Leptospira. Extracellular proteins play critical roles in the pathogenicity and survival of this pathogen in the host and environment. Extraction and analysis of extracellular proteins is a difficult task due to the abundance of enrichments like serum and bovine serum albumin in the culture medium, as is distinguishing them from the cellular proteins that may reach the analyte during extraction. In this study, extracellular proteins were separated as secretory proteins from the culture supernatant and surface proteins were separated during the washing of the cell pellet. The proteins identified were sorted based on the proportion of the cellular fractions and the extracellular fractions. The results showed the identification of 56 extracellular proteins, out of which 19 were exclusively extracellular. For those proteins, the difference in quantity with respect to their presence within the cell was found to be up to 1770-fold. Further, bioinformatics analysis elucidated characteristics and functions of the identified proteins. Orthologs of extracellular proteins in various Leptospira species were found to be closely related among different pathogenic forms. In addition to the identification of extracellular proteins, this study put forward a method for the extraction and identification of extracellular proteins.

scholarly journals A high-throughput screen for TMPRSS2 expression identifies FDA-approved compounds that can limit SARS-CoV-2 entry

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yanwen Chen ◽  
Travis B. Lear ◽  
John W. Evankovich ◽  
Mads B. Larsen ◽  
Bo Lin ◽  
...  
Keyword(s):  
Rational Design ◽  
Surface Expression ◽  
In Vitro Models ◽  
Lung Epithelial Cells ◽  
Cell Surface Receptor ◽  
Pharmacokinetic Data ◽  
Drug Induced ◽  
Global Pandemic ◽  
Surface Receptor

AbstractSARS-CoV-2 (2019-nCoV) is the pathogenic coronavirus responsible for the global pandemic of COVID-19 disease. The Spike (S) protein of SARS-CoV-2 attaches to host lung epithelial cells through the cell surface receptor ACE2, a process dependent on host proteases including TMPRSS2. Here, we identify small molecules that reduce surface expression of TMPRSS2 using a library of 2,560 FDA-approved or current clinical trial compounds. We identify homoharringtonine and halofuginone as the most attractive agents, reducing endogenous TMPRSS2 expression at sub-micromolar concentrations. These effects appear to be mediated by a drug-induced alteration in TMPRSS2 protein stability. We further demonstrate that halofuginone modulates TMPRSS2 levels through proteasomal-mediated degradation that involves the E3 ubiquitin ligase component DDB1- and CUL4-associated factor 1 (DCAF1). Finally, cells exposed to homoharringtonine and halofuginone, at concentrations of drug known to be achievable in human plasma, demonstrate marked resistance to SARS-CoV-2 infection in both live and pseudoviral in vitro models. Given the safety and pharmacokinetic data already available for the compounds identified in our screen, these results should help expedite the rational design of human clinical trials designed to combat active COVID-19 infection.

A widespread pathway for substitution of adenine by diaminopurine in phage genomes

Science ◽  
2021 ◽  
Vol 372 (6541) ◽  
pp. 512-516
Author(s):  
Yan Zhou ◽  
Xuexia Xu ◽  
Yifeng Wei ◽  
Yu Cheng ◽  
Yu Guo ◽  
...  
Keyword(s):  
Large Scale ◽  
Restriction Enzymes ◽  
Diverse Range ◽  
Host Restriction ◽  
Form And Function ◽  
Multienzyme System ◽  
Dna Modifications ◽  
Dna Production ◽  
And Function

DNA modifications vary in form and function but generally do not alter Watson-Crick base pairing. Diaminopurine (Z) is an exception because it completely replaces adenine and forms three hydrogen bonds with thymine in cyanophage S-2L genomic DNA. However, the biosynthesis, prevalence, and importance of Z genomes remain unexplored. Here, we report a multienzyme system that supports Z-genome synthesis. We identified dozens of globally widespread phages harboring such enzymes, and we further verified the Z genome in one of these phages, Acinetobacter phage SH-Ab 15497, by using liquid chromatography with ultraviolet and mass spectrometry. The Z genome endows phages with evolutionary advantages for evading the attack of host restriction enzymes, and the characterization of its biosynthetic pathway enables Z-DNA production on a large scale for a diverse range of applications.

scholarly journals Tree Diversity Reduces Fungal Endophyte Richness and Diversity in a Large-Scale Temperate Forest Experiment

Diversity ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 234 ◽  
Author(s):  
Eric A. Griffin ◽  
Joshua G. Harrison ◽  
Melissa K. McCormick ◽  
Karin T. Burghardt ◽  
John D. Parker
Keyword(s):  
Phylogenetic Diversity ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Spatial Scales ◽  
Fungal Endophytes ◽  
Fungal Endophyte ◽  
Tree Diversity ◽  
Trophic Levels ◽  
Community Diversity ◽  
And Function

Although decades of research have typically demonstrated a positive correlation between biodiversity of primary producers and associated trophic levels, the ecological drivers of this association are poorly understood. Recent evidence suggests that the plant microbiome, or the fungi and bacteria found on and inside plant hosts, may be cryptic yet important drivers of important processes, including primary production and trophic interactions. Here, using high-throughput sequencing, we characterized foliar fungal community diversity, composition, and function from 15 broadleaved tree species (N = 545) in a recently established, large-scale temperate tree diversity experiment using over 17,000 seedlings. Specifically, we tested whether increases in tree richness and phylogenetic diversity would increase fungal endophyte diversity (the “Diversity Begets Diversity” hypothesis), as well as alter community composition (the “Tree Diversity–Endophyte Community” hypothesis) and function (the “Tree Diversity–Endophyte Function” hypothesis) at different spatial scales. We demonstrated that increasing tree richness and phylogenetic diversity decreased fungal species and functional guild richness and diversity, including pathogens, saprotrophs, and parasites, within the first three years of a forest diversity experiment. These patterns were consistent at the neighborhood and tree plot scale. Our results suggest that fungal endophytes, unlike other trophic levels (e.g., herbivores as well as epiphytic bacteria), respond negatively to increasing plant diversity.

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