scholarly journals Sensing through Non-Sensing Ocular Ion Channels

2020 ◽  
Vol 21 (18) ◽  
pp. 6925
Author(s):  
Meha Kabra ◽  
Bikash Ranjan Pattnaik
Keyword(s):  
Ion Channels ◽  
Visual Processing ◽  
Signal Transmission ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Cone Dystrophy ◽  
Wide Range ◽  
The Impact

Ion channels are membrane-spanning integral proteins expressed in multiple organs, including the eye. In the eye, ion channels are involved in various physiological processes, like signal transmission and visual processing. A wide range of mutations have been reported in the corresponding genes and their interacting subunit coding genes, which contribute significantly to an array of blindness, termed ocular channelopathies. These mutations result in either a loss- or gain-of channel functions affecting the structure, assembly, trafficking, and localization of channel proteins. A dominant-negative effect is caused in a few channels formed by the assembly of several subunits that exist as homo- or heteromeric proteins. Here, we review the role of different mutations in switching a “sensing” ion channel to “non-sensing,” leading to ocular channelopathies like Leber’s congenital amaurosis 16 (LCA16), cone dystrophy, congenital stationary night blindness (CSNB), achromatopsia, bestrophinopathies, retinitis pigmentosa, etc. We also discuss the various in vitro and in vivo disease models available to investigate the impact of mutations on channel properties, to dissect the disease mechanism, and understand the pathophysiology. Innovating the potential pharmacological and therapeutic approaches and their efficient delivery to the eye for reversing a “non-sensing” channel to “sensing” would be life-changing.

Mutant p53 oncogenicity: dominant-negative or gain-of-function?

2020 ◽  
Author(s):  
Yan Stein ◽  
Ronit Aloni-Grinstein ◽  
Varda Rotter
Keyword(s):  
P53 Protein ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Mutant P53 ◽  
Gain Of Function ◽  
In Vivo Models ◽  
Oncogenic Activity ◽  
The Impact

Abstract The p53 protein is mutated in about 50% of human cancers. Aside from losing its tumor-suppressive activities, mutant p53 may acquire pro-oncogenic activity, which is facilitated by two underlying mechanisms. The first mechanism is the inhibition of co-expressed wild-type p53 (WTp53) activity, dubbed the dominant-negative effect (DNE). The second mechanism is a neomorphic pro-oncogenic activity that does not involve the inhibition of WTp53, termed gain-of-function (GOF). Throughout the years, both mechanisms were demonstrated in a plethora of in vitro and in vivo models. However, whether both account for protumorigenic activities of mutant p53 and in which contexts is still a matter of ongoing debate. Here, we discuss evidence for both DNE and GOF in a variety of models. These models suggest that both GOF and DNE can be relevant, but are highly dependent on the specific mutation type, genetic and cellular context and even the phenotype that is being assessed. In addition, we discuss how mutant and WTp53 might not exist as two separate entities, but rather as a continuum that may involve a balance between the two forms in the same cells, which could be tilted by various factors and drugs. Further elucidation of the factors that dictate the balance between the WT and mutant p53 states, as well as the factors that govern the impact of DNE and GOF in different cancer types, may lead to the development of more effective treatment regimens for cancer patients.

scholarly journals The Diagnostic Journey of a Patient with Prader–Willi-Like Syndrome and a Unique Homozygous SNURF-SNRPN Variant; Bio-Molecular Analysis and Review of the Literature

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 875
Author(s):  
Karlijn Pellikaan ◽  
Geeske M. van Woerden ◽  
Lotte Kleinendorst ◽  
Anna G. W. Rosenberg ◽  
Bernhard Horsthemke ◽  
...  
Keyword(s):  
Intellectual Disability ◽  
Single Nucleotide Polymorphism Array ◽  
Genetic Defect ◽  
Missense Variant ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Pituitary Hormone ◽  
Negative Effect

Prader–Willi syndrome (PWS) is a rare genetic condition characterized by hypotonia, intellectual disability, and hypothalamic dysfunction, causing pituitary hormone deficiencies and hyperphagia, ultimately leading to obesity. PWS is most often caused by the loss of expression of a cluster of genes on chromosome 15q11.2-13. Patients with Prader–Willi-like syndrome (PWLS) display features of the PWS phenotype without a classical PWS genetic defect. We describe a 46-year-old patient with PWLS, including hypotonia, intellectual disability, hyperphagia, and pituitary hormone deficiencies. Routine genetic tests for PWS were normal, but a homozygous missense variant NM_003097.3(SNRPN):c.193C>T, p.(Arg65Trp) was identified. Single nucleotide polymorphism array showed several large regions of homozygosity, caused by high-grade consanguinity between the parents. Our functional analysis, the ‘Pipeline for Rapid in silico, in vivo, in vitro Screening of Mutations’ (PRiSM) screen, showed that overexpression of SNRPN-p.Arg65Trp had a dominant negative effect, strongly suggesting pathogenicity. However, it could not be confirmed that the variant was responsible for the phenotype of the patient. In conclusion, we present a unique homozygous missense variant in SNURF-SNRPN in a patient with PWLS. We describe the diagnostic trajectory of this patient and the possible contributors to her phenotype in light of the current literature on the genotype–phenotype relationship in PWS.

scholarly journals In VivoEfficacy of Anuran Trypsin Inhibitory Peptides against Staphylococcal Skin Infection and the Impact of Peptide Cyclization

2015 ◽  
Vol 59 (4) ◽  
pp. 2113-2121 ◽  
Author(s):  
U. Malik ◽  
O. N. Silva ◽  
I. C. M. Fensterseifer ◽  
L. Y. Chan ◽  
R. J. Clark ◽  
...  
Keyword(s):  
Antimicrobial Agents ◽  
Cyclic Peptide ◽  
Sequence Similarity ◽  
Infection Model ◽  
Content Type ◽  
Wide Range ◽  
Inhibitory Activities ◽  
The Impact

ABSTRACTStaphylococcus aureusis a virulent pathogen that is responsible for a wide range of superficial and invasive infections. Its resistance to existing antimicrobial drugs is a global problem, and the development of novel antimicrobial agents is crucial. Antimicrobial peptides from natural resources offer potential as new treatments against staphylococcal infections. In the current study, we have examined the antimicrobial properties of peptides isolated from anuran skin secretions and cyclized synthetic analogues of these peptides. The structures of the peptides were elucidated by nuclear magnetic resonance (NMR) spectroscopy, revealing high structural and sequence similarity with each other and with sunflower trypsin inhibitor 1 (SFTI-1). SFTI-1 is an ultrastable cyclic peptide isolated from sunflower seeds that has subnanomolar trypsin inhibitory activity, and this scaffold offers pharmaceutically relevant characteristics. The five anuran peptides were nonhemolytic and noncytotoxic and had trypsin inhibitory activities similar to that of SFTI-1. They demonstrated weakin vitroinhibitory activities againstS. aureus, but several had strong antibacterial activities againstS. aureusin anin vivomurine wound infection model. pYR, an immunomodulatory peptide fromRana sevosa, was the most potent, with complete bacterial clearance at 3 mg · kg−1. Cyclization of the peptides improved their stability but was associated with a concomitant decrease in antimicrobial activity. In summary, these anuran peptides are promising as novel therapeutic agents for treating infections from a clinically resistant pathogen.

scholarly journals Cernunnos influences human immunoglobulin class switch recombination and may be associated with B cell lymphomagenesis

2012 ◽  
Vol 209 (2) ◽  
pp. 291-305 ◽  
Author(s):  
Likun Du ◽  
Roujun Peng ◽  
Andrea Björkman ◽  
Noel Filipe de Miranda ◽  
Cornelia Rosner ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
Class Switch Recombination ◽  
B Cell Lymphoma ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
End Joining ◽  
Class Switch

Cernunnos is involved in the nonhomologous end-joining (NHEJ) process during DNA double-strand break (DSB) repair. Here, we studied immunoglobulin (Ig) class switch recombination (CSR), a physiological process which relies on proper repair of the DSBs, in B cells from Cernunnos-deficient patients. The pattern of in vivo generated CSR junctions is altered in these cells, with unusually long microhomologies and a lack of direct end-joining. The CSR junctions from Cernunnos-deficient patients largely resemble those from patients lacking DNA ligase IV, Artemis, or ATM, suggesting that these factors are involved in the same end-joining pathway during CSR. By screening 269 mature B cell lymphoma biopsies, we also identified a somatic missense Cernunnos mutation in a diffuse large B cell lymphoma sample. This mutation has a dominant-negative effect on joining of a subset of DNA ends in an in vitro NHEJ assay. Translocations involving both Ig heavy chain loci and clonal-like, dynamic IgA switching activities were observed in this tumor. Collectively, our results suggest a link between defects in the Cernunnos-dependent NHEJ pathway and aberrant CSR or switch translocations during the development of B cell malignancies.

ARCD-1, an apobec-1-related cytidine deaminase, exerts a dominant negative effect on C to U RNA editing

2001 ◽  
Vol 281 (6) ◽  
pp. C1904-C1916 ◽  
Author(s):  
Shrikant Anant ◽  
Debnath Mukhopadhyay ◽  
Vakadappu Sankaranand ◽  
Susan Kennedy ◽  
Jeffrey O. Henderson ◽  
...  
Keyword(s):  
Rna Editing ◽  
Rna Binding ◽  
Cytidine Deaminase ◽  
Binding Activity ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Mrna Editing ◽  
Apob Mrna

Mammalian apolipoprotein B (apoB) C to U RNA editing is catalyzed by a multicomponent holoenzyme containing a single catalytic subunit, apobec-1. We have characterized an apobec-1 homologue, ARCD-1, located on chromosome 6p21.1, and determined its role in apoB mRNA editing. ARCD-1 mRNA is ubiquitously expressed; phylogenetic analysis reveals it to be a distant member of the RNA editing family. Recombinant ARCD-1 demonstrates cytidine deaminase and apoB RNA binding activity but does not catalyze C to U RNA editing, either in vitro or in vivo. Although not competent itself to mediate deamination of apoB mRNA, ARCD-1 inhibits apobec-1-mediated C to U RNA editing. ARCD-1 interacts and heterodimerizes with both apobec-1 and apobec-1 complementation factor (ACF) and localizes to both the nucleus and cytoplasm of transfected cells. Together, the data suggest that ARCD-1 is a novel cytidine deaminase that interacts with apobec-1 and ACF to inhibit apoB mRNA editing, possibly through interaction with other protein components of the apoB RNA editing holoenzyme.

scholarly journals Identification of a druggable protein-protein interaction site between mutant p53 and its stabilizing chaperone DNAJA1

2020 ◽  
pp. jbc.RA120.014749
Author(s):  
Xin Tong ◽  
Dandan Xu ◽  
Rama K. Mishra ◽  
Ryan D Jones ◽  
Leyu Sun ◽  
...  
Keyword(s):  
Small Molecule ◽  
Tp53 Gene ◽  
Dominant Negative ◽  
Site Directed Mutagenesis ◽  
Dominant Negative Effect ◽  
Mutant P53 ◽  
Missense Mutations ◽  
Oncogenic Function

The TP53 gene is the most frequently mutated gene in human cancers, and the majority of TP53 mutations are missense mutations. As a result, these mutant p53 (mutp53) either directly lose wild-type p53 (wtp53) tumor suppressor function or exhibit a dominant negative effect over wtp53. In addition, some mutp53 have acquired new oncogenic function (gain of function). Therefore, targeting mutp53 for its degradation, may serve as a promising strategy for cancer prevention and therapy. Based on our previous finding that farnesylated DNAJA1 is a crucial chaperone in maintaining mutp53 stabilization, and by using an in silico approach, we built 3-D homology models of human DNAJA1 and mutp53R175H proteins, identified the interacting pocket in the DNAJA1-mutp53R175H complex, and found one critical druggable small molecule binding  site in the DNAJA1 glycine/phenylalanine rich region. We confirmed that the interacting pocket in the DNAJA1-mutp53R175H complex was crucial for stabilizing mutp53R175H using a site-directed mutagenesis approach. We further screened a drug-like library to identify a promising small molecule hit (GY1-22) against the interacting pocket in DNAJA1-mutp53R175H complex. The GY1-22 compound displayed an effective activity against DNAJA1-mutp53R175H complex. Treatment with GY1-22 significantly reduced mutp53 protein levels, enhanced Waf1p21 expression, suppressed cyclin D1 expression, and inhibited mutp53-driven pancreatic cancer growth both in vitro and in vivo. Together, our results indicate that the interacting pocket in the DNAJA1-mutp53R175H complex is critical for mutp53’s stability and oncogenic function, and DNAJA1 is a robust therapeutic target for developing the efficient small molecule inhibitors against oncogenic mutp53.

Photopatterned Membranes and Chemical Gradients Enable Scalable Phenotypic Organization of Primary Human Colon Epithelial Models

2019 ◽  
Author(s):  
Sam Hinman ◽  
Yuli Wang ◽  
Nancy Allbritton
Keyword(s):  
Human Colon ◽  
Cell Types ◽  
Self Renewal ◽  
Chemical Gradients ◽  
Plate Spacing ◽  
Wide Range ◽  
Cell Compartmentalization ◽  
The Impact

Biochemical gradients across the intestinal epithelium play a major role in governing intestinal stem cell compartmentalization, differentiation dynamics, and organ-level self-renewal. Advances in primary cell-derived <i>in vitro</i> models, in which a full suite of stem and differentiated cell types are present, have vastly accelerated our understanding of intestinal homeostasis and disease. However, scalable platforms that recapitulate the architecture and gradients present <i>in vivo</i> are absent. We present a platform in which individually addressable arrays of chemical gradients along the crypt long axis can be generated, enabling scalable culture of <i>in vitro</i> colonic epithelial replicas. The platform utilizes standardized well plate spacing, maintains access to basal and luminal compartments, and relies on a photopatterned porous membrane to act as diffusion windows while supporting the<i> in vitro </i>crypts. Simultaneous fabrication of 3,875 crypts over a single membrane was developed. Growth factor gradients were modelled and then experimentally optimized to promote long-term health and self-renewal of the crypts which were assayed <i>in situ</i> by confocal fluorescence microscopy. The cultured <i>in vitro</i> crypt arrays successfully recapitulated the architecture, stem/proliferative and differentiated cell compartmentalization, and luminal-to-basal polarity observed <i>in vivo</i>. Furthermore, known signaling regulators produced measurable and predictable effects on the proliferative and differentiated cell compartments. This platform is readily adaptable to the screening of tissue from individual patients to assay the impact of food and bacterial metabolites and/or drugs on colonic crypt dynamics. Importantly, the cassette is compatible with a wide range of sensing/detection modalities, and the developed fabrication methods should find applications for other cell and tissue types.

scholarly journals Inhibition of proteolysis and cell cycle progression in a multiubiquitination-deficient yeast mutant.

1994 ◽  
Vol 14 (8) ◽  
pp. 5501-5509 ◽  
Author(s):  
D Finley ◽  
S Sadis ◽  
B P Monia ◽  
P Boucher ◽  
D J Ecker ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Turnover ◽  
Cell Cycle Progression ◽  
Critical Role ◽  
Sole Source ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Polyubiquitin Gene

The degradation of many proteins requires their prior attachment to ubiquitin. Proteolytic substrates are characteristically multiubiquitinated through the formation of ubiquitin-ubiquitin linkages. Lys-48 of ubiquitin can serve as a linkage site in the formation of such chains and is required for the degradation of some substrates of this pathway in vitro. We have characterized the recessive and dominant effects of a Lys-48-to-Arg mutant of ubiquitin (UbK48R) in Saccharomyces cerevisiae. Although UbK48R is expected to terminate the growth of Lys-48 multiubiquitin chains and thus to exert a dominant negative effect on protein turnover, overproduction of UbK48R in wild-type cells results in only a weak inhibition of protein turnover, apparently because the mutant ubiquitin can be removed from multiubiquitin chains. Surprisingly, expression of UbK48R complements several phenotypes of polyubiquitin gene (UB14) deletion mutants. However, UbK48R cannot serve as a sole source of ubiquitin in S. cerevisiae, as evidenced by its inability to rescue the growth of ubi1 ubi2 ubi3 ubi4 quadruple mutants. When provided solely with UbK48R, cells undergo cell cycle arrest with a terminal phenotype characterized by replicated DNA, mitotic spindles, and two-lobed nuclei. Under these conditions, degradation of amino acid analog-containing proteins is severely inhibited. Thus, multiubiquitin chains containing Lys-48 linkages play a critical role in protein degradation in vivo.

scholarly journals A dominant-negative variant in the dopamine transporter PDZ-binding motif is linked to parkinsonism and neuropsychiatric disease

2020 ◽  
Author(s):  
Freja Herborg ◽  
Kathrine L. Jensen ◽  
Sasha Tolstoy ◽  
Natascha V. Arends ◽  
Leonie P. Posselt ◽  
...  
Keyword(s):  
Dopamine Transporter ◽  
Early Onset ◽  
Surface Expression ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Binding Motif ◽  
Missense Mutations ◽  
Neuropsychiatric Disease

AbstractDopaminergic dysfunction is central to movement disorders and mental diseases. The dopamine transporter (DAT) is essential for the regulation of extracellular dopamine but the genetic and mechanistic link between DAT function and dopamine-related pathologies remains elusive. Particularly, the pathophysiological significance of monoallelic missense mutations in DAT is unknown. Here we identify a novel coding DAT variant, DAT-K619N, in a patient with early-onset parkinsonism and comorbid neuropsychiatric disease and in 22 individuals from exome-sequenced samples of neuropsychiatric patients. The variant localizes to the critical C-terminal PDZ-binding motif of DAT and causes reduced uptake capacity, decreased surface expression, and accelerated turnover of DAT in vitro. In vivo, we demonstrate that expression of DAT-K619N in mice and dropsophila imposes impairments in dopamine transmission with accompanying changes in dopamine-directed behaviors. Importantly, both cellular studies and viral overexpression of DAT-K619N in mice show that DAT-K619N has a dominant-negative effect which collectively implies that a single dominant-negative genetic DAT variant can confer risk for neuropsychiatric disease and neurodegenerative early-onset parkinsonism.

scholarly journals Immunotoxic Effects Induced by Microcystins and Cylindrospermopsin: A Review

Toxins ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 711
Author(s):  
Leticia Diez-Quijada ◽  
Maria del Monte Benítez-González ◽  
María Puerto ◽  
Angeles Jos ◽  
Ana M. Cameán
Keyword(s):  
Immune System ◽  
Scientific Literature ◽  
Anthropogenic Activities ◽  
In Vivo Studies ◽  
Wide Range ◽  
Full Picture ◽  
Underlying Mechanisms ◽  
The Impact

Cyanotoxin occurrence is gaining importance due to anthropogenic activities, climate change and eutrophication. Among them, Microcystins (MCs) and Cylindrospermopsin (CYN) are the most frequently studied due to their ubiquity and toxicity. Although MCs are primary classified as hepatotoxins and CYN as a cytotoxin, they have been shown to induce deleterious effects in a wide range of organs. However, their effects on the immune system are as yet scarcely investigated. Thus, to know the impact of cyanotoxins on the immune system, due to its importance in organisms’ homeostasis, is considered of interest. A review of the scientific literature dealing with the immunotoxicity of MCs and CYN has been performed, and both in vitro and in vivo studies have been considered. Results have confirmed the scarcity of reports on the topic, particularly for CYN. Decreased cell viability, apoptosis or altered functions of immune cells, and changed levels and mRNA expression of cytokines are among the most common effects reported. Underlying mechanisms, however, are still not yet fully elucidated. Further research is needed in order to have a full picture of cyanotoxin immunotoxicity.

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