Myelin basic protein (MBP) is the second most abundant protein in the central nervous system and is responsible for structural maintenance of the myelin sheath covering axons. Previously, we showed ...that MBP has a more proactive role in the oligodendrocyte homeostasis, interacting with membrane-associated proteins, including integral membrane protein 2B (ITM2B or Bri2) that is associated with familial dementias. Here, we report that the molecular dynamics of the in silico-generated MBP-Bri2 complex revealed that MBP covers a significant portion of the Bri2 ectodomain, assumingly trapping the furin cleavage site, while the surface of the BRICHOS domain, which is responsible for the multimerization and activation of the Bri2 high-molecular-weight oligomer chaperone function, remains unmasked. These observations were supported by the co-expression of MBP with Bri2, its mature form, and disease-associated mutants, which showed that in mammalian cells, MBP indeed modulates the post-translational processing of Bri2 by restriction of the furin-catalyzed release of its C-terminal peptide. Moreover, we showed that the co-expression of MBP and Bri2 also leads to an altered cellular localization of Bri2, restricting its membrane trafficking independently of the MBP-mediated suppression of the Bri2 C-terminal peptide release. Further investigations should elucidate if these observations have physiological meaning in terms of Bri2 as a MBP chaperone activated by the MBP-dependent postponement of Bri2 membrane trafficking.
Myelin basic protein (MBP) is one of the key proteins in the development of multiple sclerosis (MS). However, very few intracellular MBP partners have been identified up to now. In order to find ...proteins interacting with MBP in the brain, an expression library from the human brain was screened using a yeast two-hybrid system. Here we showed that MBP interacts with the C-terminal 24-residue peptide of Integral transmembrane protein II associated with familial British and Danish dementia (ITM2B/Bri2 or Bri2). This peptide (Bri23R) was one residue longer than the known Bri23 peptide, which is cleaved from the C-terminus of Bri2 during its maturation in the Golgi and has physiological activity as a modulator of amyloid precursor protein processing. Since the spatial structures for both MBP and Bri2 were not known, we used computational methods of structural biology including an artificial intelligence system AlphaFold2 and high ambiguity driven protein-protein docking (HADDOCK 2.1) to gain a mechanistic explanation of the found protein-protein interaction and elucidate a possible structure of the complex of MBP with Bri23R peptide. As expected, MBP was mostly unstructured, although it has well-defined α-helical regions, while Bri23R forms a stable β-hairpin. Simulation of the interaction between MBP and Bri23R in two different environments, as parts of the two-hybrid system fusion proteins and in the form of single polypeptides, showed that MBP twists around Bri23R. The observed interaction results in the adjustment of the size of the internal space between MBP α-helices to the size of the β-hairpin of Bri23R. Since Bri23 is known to inhibit aggregation of amyloid oligomers, and the association of MBP to the inner leaflet of the membrane bilayer shares features with amyloid fibril formation, Bri23 may serve as a peptide chaperon for MBP, thus participating in myelin membrane assembly.
Starting from a screening hit, a set of analogs was synthesized based on a 4-(2-aminoethyl)piperidine core not associated previously with trace amine-associated receptor 1 (TAAR1) modulation in the ...literature. Several structure–activity relationship generalizations have been drawn from the observed data, some of which were corroborated by molecular modeling against the crystal structure of TAAR1. The four most active compounds (EC50 for TAAR1 agonistic activity ranging from 0.033 to 0.112 μM) were nominated for evaluation in vivo. The dopamine transporter knockout (DAT-KO) rat model of dopamine-dependent hyperlocomotion was used to evaluate compounds’ efficacy in vivo. Out of four compounds, only one compound (AP163) displayed a statistically significant and dose-dependent reduction in hyperlocomotion in DAT-KO rats. As such, compound AP163 represents a viable lead for further preclinical characterization as a potential novel treatment option for disorders associated with increased dopaminergic function, such as schizophrenia.
Myelin basic protein (MBP) is one of the key structural elements of the myelin sheath and has autoantigenic properties in multiple sclerosis (MS). Its intracellular interaction network is still ...partially deconvoluted due to the unfolded structure, abnormally basic charge, and specific cellular localization. Here we used the fusion protein of MBP with TurboID, an engineered biotin ligase that uses ATP to convert biotin to reactive biotin-AMP that covalently attaches to nearby proteins, to determine MBP interactome. Despite evident benefits, the proximity labeling proteomics technique generates high background noise, especially in the case of proteins tending to semi-specific interactions. In order to recognize unique MBP partners, we additionally mapped protein interaction networks for deaminated MBP variant and cyclin-dependent kinase inhibitor 1 (p21), mimicking MBP in terms of natively unfolded state, size and basic amino acid clusters. We found that in the plasma membrane region, MBP is colocalized with adhesion proteins occludin and myelin protein zero-like protein 1, solute carrier family transporters ZIP6 and SNAT1, Eph receptors ligand Ephrin-B1, and structural components of the vesicle transport machinery-synaptosomal-associated protein 23 (
), vesicle-associated membrane protein 3 (
), protein transport protein hSec23B and cytoplasmic dynein 1 heavy chain 1. We also detected that MBP potentially interacts with proteins involved in Fe
and lipid metabolism, namely, ganglioside GM2 activator protein, long-chain-fatty-acid-CoA ligase 4 (
), NADH-cytochrome b5 reductase 1 (
) and metalloreductase STEAP3. Assuming the emerging role of ferroptosis and vesicle cargo docking in the development of autoimmune neurodegeneration, MBP may recruit and regulate the activity of these processes, thus, having a more inclusive role in the integrity of the myelin sheath.
Using the envelope glycoprotein B (gB) crystal structure and digital prediction algorithm, the B- and T-cell antigenic determinants (epitopes) of human herpesvirus 1 (HHV-1), also known as herpes ...simplex virus 1 (HSV-1), were generated, and the method for their production in the form of recombinant proteins was proposed. First, the structure of the surface topological domain (ectodomain or spike) of gB with mapped epitopes was analyzed, and the most stable and immunogenic (due to their enrichment with B-and T-cell epitopes) subdomains were selected for the modeling of subunit vaccine prototypes using the AlphaFold2 (Google DeepMind, London, UK) artificial intelligence system. The proposed candidate vaccines included both small (about 100 amino acids) monomeric polypeptides, which were ideal for recombinant expression as fusion proteins, and a more complex polypeptide, which, due to its trimeric fold, looks like a miniature analog of the gB ectodomain. In this miniature analog, the ectodomain regions with the potential to interfere efficacious expression of soluble recombinant protein in Escherichia coli have been removed. The structural stability of the modeled proteins, confirmed by molecular dynamics simulation and host immune responses, predicted in silico, indicates the suitability of the two suggested polypeptides for generating subunit vaccines using recombinant DNA technology.
Intrinsically disordered myelin basic protein (MBP) is one of the key autoantigens in autoimmune neurodegeneration and multiple sclerosis particularly. MBP is highly positively charged and lacks ...distinct structure in solution and therefore its intracellular partners are still mostly enigmatic. Here we used combination of formaldehyde-induced cross-linking followed by immunoprecipitation and liquid chromatography-tandem mass spectrometry (LC-MS/MS) to elucidate the interaction network of MBP in mammalian cells and provide the list of potential MBP interacting proteins. Our data suggest that the largest group of MBP-interacting proteins belongs to cellular proteins involved in the protein translation machinery, as well as in the spatial and temporal regulation of translation. MBP interacts with core ribosomal proteins, RNA helicase Ddx28 and RNA-binding proteins STAU1, TDP-43, ADAR-1 and hnRNP A0, which are involved in various stages of RNA biogenesis and processing, including specific maintaining MBP-coding mRNA. Among MBP partners we identified CTNND1, which has previously been shown to be necessary for myelinating Schwann cells for cell-cell interactions and the formation of a normal myelin sheath. MBP binds proteins MAGEB2/D2 associated with neurotrophin receptor p75NTR, involved in pathways that promote neuronal survival and neuronal death. Finally, we observed that MBP interacts with RNF40-a component of heterotetrameric Rnf40/Rnf20 E3 ligase complex, recruited by Egr2, which is the central transcriptional regulator of peripheral myelination. Concluding, our data suggest that MBP may be more actively involved in myelination not only as a main building block but also as a self-regulating element.
Trace amines are a group of biogenic amines that are structurally and functionally close to classical monoamine neurotransmitters. Trace amine-associated receptors (TAARs) are emerging as promising ...targets for treating neuropsychiatric disorders. It has been documented that all TAARs, apart from TAAR1, function as olfactory receptors involved in sensing innate odors encoded by volatile amines. However, recently, brain expression and function of TAAR5 were also demonstrated. In this study, we assessed the behavior, brain neurochemistry, and electrophysiology changes in knock-out mice lacking Trace amine-associated receptor 2 (TAAR2) but expressing beta-Galactosidase mapping expression of TAAR2 receptors. As expected, we detected beta-Galactosidase staining in the glomerular layer of the olfactory bulb. However, we also found staining in the deeper layers of the olfactory bulb and several brain regions, including the hippocampus, cerebellum, cortex, raphe nuclei, hypothalamus, and habenula, indicating that TAAR2 receptors are not only expressed in the olfactory system but are also present in the limbic brain areas that receive olfactory input. In behavioral experiments, TAAR2 knock-out (TAAR2-KO) mice showed increased locomotor activity and less immobility in the forced swim test, with no changes in anxiety level. Furthermore, TAAR2-KO mice showed alterations in brain electrophysiological activity-particularly, decreased spectral power of the cortex and striatum in the 0, 9-20 Hz range. TAAR2-KO mice also had elevated tissue dopamine levels in the striatum and an increased dopaminergic neuron number in the Substantia Nigra. In addition, an increased brain-derived neurotrophic factor (BDNF) mRNA level in the striatum and Monoamine Oxidase B (MAO-B) mRNA level in the striatum and midbrain was found in TAAR2-KO mice. Importantly, TAAR2-KO mice demonstrated an increased neuroblast-like and proliferating cell number in the subventricular and subgranular zone, indicating increased adult neurogenesis. These data indicate that in addition to its role in the innate olfaction of volatile amines, TAAR2 is expressed in limbic brain areas and regulates the brain dopamine system, neuronal electrophysiological activity, and adult neurogenesis. These findings further corroborated observations in TAAR1-KO and TAAR5-KO mice, indicating common for TAAR family pattern of expression in limbic brain areas and role in regulating monoamine levels and adult neurogenesis, but with variable involvement of each subtype of TAAR receptors in these functions.
The properties of the root system are especially important under drought, which is currently the most significant climate threat worldwide. The aim of this work was to investigate the relationship ...between the root system development and vernalization requirements under contrasting irrigation conditions in bread wheat using lines with alien introgressions. A set of single chromosome substitution lines Chinese Spring (Synthetic 6x, AABBDD) and D-genome derived introgression lines were used to study the relationships. It was found that 1A substitution resulted in a substantial reduction of root length and weight while 5D substitution, on the contrary, led to a significant increase compared to the recipient and the donor under both irrigation regimes. The greatest root weight, root length, and number of days to flowering were detected in the lines carrying a common introgression fragment of the 5D chromosome connected with the molecular marker
Xgwm292
and the
Vrn-D1
gene. The lines with the introgression in this region had a significantly larger root length and weight compared with the recipient and donor under all conditions of irrigation. The combination of prolonged vernalization and drought negatively affected the development of roots in all lines with the most pronounced effect on root weight. With the use of monosomic analysis, it was shown that the
Vrn-A1
gene has a substantial reducing effect on the root length and weight. The revealed relationship between the flowering time and root system development can be used for breeding of wheat cultivars more adapted for growing in arid conditions.
Introduction. Integration processes in higher education have recently come into particular prominence. A special place in this context is occupied by the issues of integration in the field of higher ...education of the countries of the Eurasian Economic Union, which have not yet been sufficiently reflected in research. The goal of this paper is to develop theoretical and methodological foundations for assessing the effectiveness of formation and functioning of the integration complex among the Eurasian Economic Union universities as a factor in increasing their competitiveness.
Materials and Methods. Methods of theoretical and comparative analysis, synthesis and classification, method of constructing an economic and mathematical model, method of expert assessment were used in the study.
Results. For the purpose of theoretical substantiation of one of the main forms of integration in the sphere of higher education the concept of “integration complex of universities” was introduced and its interpretation was proposed. Possible strategies for the formation of integration complexes of universities have been identified. A methodology for assessing the effectiveness of formation and functioning of the integration complex among Eurasian Economic Union universities has been developed and its integral indicator has been determined.
Discussion and Conclusion. The obtained results of the research contribute to the development of the conceptual foundations of educational integration and methodological approaches to its assessment. The materials of this article will be useful to the staff of government agencies and academic institutions that study interstate integrative processes and their effectiveness in education.
Pigment Epithelium-Derived Factor (PEDF) is a secreted glycoprotein belonging to the family of non-inhibitory serpins. It is known, that in cases of complicated myopia, the content of PEDF in aqueous ...humor of the anterior chamber is significantly reduced. Here we examined a bulk of Tenon's capsule samples obtained from various groups of myopes, to examine PEDF processing in progressive myopia. We have analyzed the distribution of full length PEDF50 and its truncated form PEDF45 in the soluble and insoluble fractions extracted from Tenon's capsule of myopic and control (non-myopic) patients using SDS-polyacrylamide gel electrophoresis, as well as monitored the proteolytic degradation of PEDF ex vivo by enzyme-linked immunosorbent assay. These results were complemented by PEDF mRNA analysis in correspondent tissues by using qPCR and immunohistochemistry analysis of PEDF distribution in normal and myopic specimens. We found that in the Tenon's capsule of patients suffering from a high myopia the level of “soluble” 45 kDa PEDF reduced by 2-fold, while the content of “insoluble” 50 kDa form of PEDF was increased by 4-fold compared to controls. Excessive amount of PEDF50 in myopic specimens have been shown to correlate with the abrogated PEDF processing rather than with an increase of its expression. Moreover, immunohistochemical staining of the myopic Tenon's capsule tissue sections revealed the halo of deposited PEDF50 in the fibroblast extracellular space. These findings suggest that in myopia limited proteolysis of PEDF is altered or abrogated. Accumulation of full-length PEDF insoluble aggregates in the fibroblast intercellular space may affect cell survival and consequently causes the destructive changes in the extracellular matrix of the eye connective tissues. As a result, the abrogation of full-length PEDF normal processing can be an important mechanism leading to biomechanical destabilization of the scleral capsule and myopia progression.
•Amount of soluble PEDF was decreased in Tenon's capsule of myopic patients.•Predominant uncleaved form of PEDF was found in Tenon's capsule of myopic patients нера≿♯епленная форма PEDF.•Uncleaved PEDF accumulates as amyloid-like fibrils in the intercellular space.
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