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Functions of about 10% of all the proteins and their associations with diseases are poorly annotated or not annotated at all. Among these proteins, there is a group of uncharacterized chromosome-specific open-reading frame genes (CxORFx) from the 'Tdark' category. The aim of the work was to reveal associations of CxORFx gene expression and ORF proteins' subinteractomes with cancer-driven cellular processes and molecular pathways. We performed systems biology and bioinformatic analysis of 219 differentially expressed CxORFx genes in cancers, an estimation of prognostic significance of novel transcriptomic signatures and analysis of subinteractome composition using several web servers (GEPIA2, KMplotter, ROC-plotter, TIMER, cBioPortal, DepMap, EnrichR, PepPSy, cProSite, WebGestalt, CancerGeneNet, PathwAX II and FunCoup). The subinteractome of each ORF protein was revealed using ten different data sources on physical protein–protein interactions (PPIs) to obtain representative datasets for the exploration of possible cellular functions of ORF proteins through a spectrum of neighboring annotated protein partners. A total of 42 out of 219 presumably cancer-associated ORF proteins and 30 cancer-dependent binary PPIs were found. Additionally, a bibliometric analysis of 204 publications allowed us to retrieve biomedical terms related to ORF genes. In spite of recent progress in functional studies of ORF genes, the current investigations aim at finding out the prognostic value of CxORFx expression patterns in cancers. The results obtained expand the understanding of the possible functions of the poorly annotated CxORFx in the cancer context. [ABSTRACT FROM AUTHOR]

Favism is a hemolytic disease due to the ingestion of fava beans in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency. There is wide inter- and intra-individual variability in the development of hemolytic crisis, and several factors influence it: quantity, quality, ripeness of fava beans, and age of onset. In this narrative review of case reports and case series, we reported the predisposing factors and clinical features for four different age groups classified as follows: pregnant women and infants (i.e., exclusively breastfed children); children, from weaned to 11 years; preadolescents and adolescents, from 11 to 18 years; and adults (18 years and older). Some symptoms developed only in specific age groups: death in infants; visual impairment in children; systolic murmur in infants, children, and adolescents; and renal failure in adults. In youngest children or pregnant women the severity is the highest. Some other symptoms were present in all: jaundice, increased bilirubin, splenomegaly, hepatomegaly, discolored urine, tachycardia, pallor, abdominal pain, malaise, vomit, nausea, and dizziness. Laboratory findings are characterized by anemia, reticulocytosis, elevated bilirubin level, and sometimes urinary urobilinogen and methemoglobinemia. In most cases the symptomatology is self-limited and does not release sequelae, but hospitalization and transfusion are often required. [ABSTRACT FROM AUTHOR]

Background: Enterovirus A71 (EV-A71) is a neurotropic virus which may cause severe neural complications, especially in infants and children. The clinical manifestations include hand-foot-and-mouth disease, herpangina, brainstem encephalitis, pulmonary edema, and other severe neurological diseases. Although there are some vaccines approved, the post-marketing surveillance is still unavailable. In addition, there is no antiviral drugs against EV-A71 available. Methods: In this study, we identified a novel antibody that could inhibit viral growth through a human single chain variable fragment (scFv) library expressed in mammalian cells and panned by infection with lethal dose of EV-A71. Results: We identified that the host protein α-enolase (ENO1) is the target of this scFv, and anti-ENO1 antibody was found to be more in mild cases than severe EV-A71 cases. Furthermore, we examined the antiviral activity in a mouse model. We found that the treatment of the identified 07-human IgG1 antibody increased the survival rate after virus challenge, and significantly decreased the viral RNA and the level of neural pathology in brain tissue. Conclusions: Collectively, through a promising intracellular scFv library expression and screening system, we found a potential scFv/antibody which targets host protein ENO1 and can interfere with the infection of EV-A71. The results indicate that the usage and application of this antibody may offer a potential treatment against EV-A71 infection. [ABSTRACT FROM AUTHOR]

The presence of many completely uncharacterized proteins, even in well-studied organisms such as humans, seriously hampers a full understanding of the functioning of living cells. One such example is the human protein C12ORF4, which belongs to the DUF2362 family, present in many eukaryotic lineages and conserved in metazoans. The only functional information available on C12ORF4 (Chromosome 12 Open Reading Frame 4) is its involvement in mast cell degranulation and its being a genetic cause of autosomal intellectual disability. Bioinformatics analysis of the DUF2362 family provides strong evidence that it is a novel member of the Macro clan/superfamily. Sequence similarity analysis versus other representatives of the Macro superfamily of ADP-ribose-binding proteins and mapping sequence conservation on predicted three-dimensional structure provides hypotheses regarding the molecular function for members of the DUF2362 family. For example, the available functional data suggest a possible role for C12ORF4 in ADP-ribosylation signaling in asthma and related inflammatory diseases. This novel family appears to be a likely novel ADP-ribosylation “reader” and “eraser,” a previously unnoticed putative new player in cell signaling by this emerging post-translational modification. [ABSTRACT FROM AUTHOR]

The high specificity of antibodies for their antigen allows a fine discrimination of target conformations and post-translational modifications, making antibodies the first choice tool to interrogate the proteome. We describe here an approach based on a large-scale intracellular expression and selection of antibody fragments in eukaryotic cells, so-called intrabodies, and the subsequent identification of their natural target within living cell. Starting from a phenotypic trait, this integrated system allows the identification of new therapeutic targets together with their companion inhibitory intrabody. We applied this system in a model of allergy and inflammation. We first cloned a large and highly diverse intrabody library both in a plasmid and a retroviral eukaryotic expression vector. After transfection in the RBL-2H3 rat basophilic leukemia cell line, we performed seven rounds of selection to isolate cells displaying a defect in FcεRI-induced degranulation. We used high throughput sequencing to identify intrabody sequences enriched during the course of selection. Only one intrabody was common to both plasmid and retroviral selections, and was used to capture and identify its target from cell extracts. Mass spectrometry analysis identified protein RGD1311164 (C12orf4), with no previously described function. Our data demonstrate that RGD1311164 is a cytoplasmic protein implicated in the early signaling events following FcεRI-induced cell activation. This work illustrates the strength of the intrabody-based in-cell selection, which allowed the identification of a new player in mast cell activation together with its specific inhibitor intrabody. [ABSTRACT FROM AUTHOR]

Presents the table of contents for the December 1996 issue of the periodical "Clinical & Experimental Immunology."

Some patients with chronic hepatitis C develop liver/kidney microsome type 1 antibodies. Some of these autoantibodies are directed against the cytochrome P450IID6. The aim of this study was to characterize the immunogenic sites on the P450IID6 molecule recognized by autoantibodies from individuals infected with the hepatitis C virus. Serum from 24 patients with markers of hepatitis C infection and liver/kidney microsome type 1 antibodies were tested by immunoblotting with human liver microsomal proteins. Eight of them with anti-P450IID6 antibodies were tested with synthetic peptides representing P450IID6 putative immunogenic sites and for their capacity to inhibit in vitro P450IID6 enzymatic activity. Anti-P450IID6 antibodies in the sera of chronic hepatitis C patients were of IgG subclass 1 and in one patient also of IgM type. These sera recognized different P450IID6 peptide sequences consisting of amino acids (aa) 200-214 and aa 271-339. The synthetic peptide between aa 321 and 339 appears to represent the main antigenic site. Five out of eight anti-P450IID6 positive sera were also able to inhibit P450IID6 enzymatic activity in vitro at a dilution of 1:100. The anti-P450IID6 autoimmune response in chronic hepatitis C patients is polyclonal, probably inducible, and maintained by the liberation of P450IID6 by hepatocyte lysis. The characterization of the P450IID6 immunogenic site recognized by patients with hepatitis C infection may enable a specific test to be designed to identify those patients with autoimmune hepatitis type 2. [ABSTRACT FROM AUTHOR]

To date very few promising leads from natural products (NP) secondary metabolites with antiviral and immunomodulatory properties have been identified for promising/potential intervention for COVID-19. Using in-silico docking studies and genome based various molecular targets, and their in vitro anti-SARS CoV-2 activities against whole cell and/or selected protein targets, we select a few compounds of interest, which can be used as potential leads to counteract effects of uncontrolled innate immune responses, in particular those related to the cytokine storm. A critical factor for prevention and treatment of SARS-CoV-2 infection relates to factors independent of viral infection or host response. They include population-related variables such as concurrent comorbidities and genetic factors critically relevant to COVID-19 health disparities. We discuss population risk factors related to SARS-CoV-2. In addition, we focus on virulence related to glucose-6-phosphate dehydrogenase deficiency (G6PDd), the most common human enzymopathy. Review of data on the response of individuals and communities with high prevalence of G6PDd to NP, prompts us to propose the rationale for a population-specific management approach to rationalize design of therapeutic interventions of SARS-CoV-2 infection, based on use of NP. This strategy may lead to personalized approaches and improve disease-related outcomes. [ABSTRACT FROM AUTHOR]

In biomedical research, there is an ongoing demand for new technologies to elucidate disease mechanisms and develop novel therapeutics. This requires comprehensive understanding of cellular processes and their pathophysiology based on reliable information on abundance, localization, post-translational modifications and dynamic interactions of cellular components. Traceable intracellular binding molecules provide new opportunities for real-time cellular diagnostics. Most prominently, intrabodies derived from antibody fragments of heavy-chain only antibodies of camelids (nanobodies) have emerged as highly versatile and attractive probes to study and manipulate antigens within the context of living cells. In this review, we provide an overview on the selection, delivery and usage of intrabodies to visualize and monitor cellular antigens in living cells and organisms. Additionally, we summarize recent advances in the development of intrabodies as cellular biosensors and their application to manipulate disease-related cellular processes. Finally, we highlight switchable intrabodies, which open entirely new possibilities for real-time cell-based diagnostics including live-cell imaging, target validation and generation of precisely controllable binding reagents for future therapeutic applications. [ABSTRACT FROM AUTHOR]

Background: Intellectual disability (ID) is a feature of many rare diseases caused by thousands of genes. This genetic heterogeneity implies that pathogenic variants in a specific gene are found only in a small number of patients, and difficulties arise in the definition of prevailing genotype and characteristic phenotype associated with that gene. One of such very rare disorders is autosomal recessive ID type 66 (OMIM #618221) caused by defects in C12orf4. Up to now, six families have been reported with mostly truncating variants. The spectrum of the clinical phenotype was not emphasized in previous reports, and detailed phenotype was not always available from previous patients, especially from large cohort studies. Methods: Exome sequencing was performed in a consanguineous Armenian family with two affected adult brothers. Results: The patients carry a novel homozygous nonsense C12orf4 variant. The integration of previous data and phenotyping of the brothers indicate that the clinical picture of C12orf4 defects involves hypotonia in infancy, rather severe ID, speech impairment, and behavioral problems such as aggressiveness, unstable mood, and autistic features. Several other symptoms are more variable and less consistent. Conclusion: This rather nonsyndromic and nonspecific clinical picture implies that additional patients with C12orf4 defects will likely continue to be identified using the "genotype‐first" approach, rather than based on clinical assessment. The phenotype needs further delineation in future reports. [ABSTRACT FROM AUTHOR]

Background: Doxycycline is an antibiotic used in combination with quinine or artesunate for malaria treatment or alone for malaria chemoprophylaxis. Recently, one prophylactic failure has been reported, and several studies have highlighted in vitro doxycycline decreased susceptibility in Plasmodium falciparum isolates from different areas. The genetic markers that contribute to detecting and monitoring the susceptibility of P. falciparum to doxycycline, the pfmdt and pftetQ genes, have recently been identified. However, these markers are not sufficient to explain in vitro decreased susceptibility of P. falciparum to doxycycline. In this paper, the association between polymorphism of the small sub-unit ribosomal RNA apicoplastic gene pfssrRNA (PFC10_API0057) and in vitro susceptibilities of P. falciparum isolates to doxycycline were investigated., Methods: Doxycycline IC50 determinations using the hypoxanthine uptake inhibition assay were performed on 178 African and Thai P. falciparum isolates. The polymorphism of pfssrRNA was investigated in these samples by standard PCR followed by sequencing., Results: No point mutations were found in pfssrRNA in the Thai or African isolates, regardless of the determined IC50 values., Conclusions: The pfssrRNA gene is not associated with in vitro decreased susceptibility of P. falciparum to doxycycline. Identifying new in vitro molecular markers associated with reduced susceptibility is needed, to survey the emergence of doxycycline resistance.

Background: The objective of this study was to evaluate the distribution of a series of independent doxycycline inhibitory concentration 50% (IC50) values to validate the trimodal distribution previously described and to validate the use of the pftetQ and pfmdt genes as molecular markers of decreased in vitro doxycycline susceptibility in Plasmodium falciparum malaria., Methods: Doxycycline IC50 values, from 484 isolates obtained at the French National Reference Centre for Imported Malaria (Paris) between January 2006 and December 2010, were analysed for the first time by a Bayesian mixture modelling approach to distinguish the different in vitro phenotypic groups by their IC50 values. Quantitative real-time polymerase chain reaction was used to evaluate the pftetQ and pfmdt copy numbers of 89 African P. falciparum isolates that were randomly chosen from the phenotypic groups., Results: The existence of at least three doxycycline phenotypes was demonstrated. The mean doxycycline IC50 was significantly higher in the group with a pftetQ copy number >1 compared to the group with a pftetQ copy number = 1 (33.17 μM versus 17.23 μM) and the group with a pfmdt copy number >1 (28.28 μM versus 16.11 μM). There was a significant difference between the combined low and medium doxycycline IC50 group and the high IC50 group in terms of the per cent of isolates with one or more copy numbers of the pftetQ gene (0% versus 20.69%) or pfmdt gene (8.33% versus 37.93%). In the logistic regression model, the pfmdt and pftetQ copy numbers >1 (odds ratio = 4.65 and 11.47) were independently associated with the high IC50 group., Conclusions: Copy numbers of pftetQ and pfmdt are potential predictive molecular markers of decreased susceptibility to doxycycline.

Presents the table of contents for the November 1996 issue of the journal "Clinical and Experimental Immunology."