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Neurology Jul 2024Obesity is hypothesized to induce a hypercoagulable state that increases stroke risk. The molecular mechanisms underlying this association are largely uncharacterized....
BACKGROUND AND OBJECTIVES
Obesity is hypothesized to induce a hypercoagulable state that increases stroke risk. The molecular mechanisms underlying this association are largely uncharacterized. We aimed to apply mendelian randomization to identify whether the association of genetically proxied body mass index (BMI) with cardioembolic stroke risk is mediated by changes in levels of circulating coagulation factors.
METHODS
Genetic proxies for BMI and levels of circulating coagulation factors were obtained, respectively, from the Genetic Investigation of ANthropometric Traits consortium (n = 694,649) and deCODE cohort (n = 35,559). Genetic associations with cardioembolic stroke risk were obtained from the GIGASTROKE consortium (10,804 cases and 1,234,804 controls). We performed a two-sample mendelian randomization analysis testing the association of genetically proxied BMI with cardioembolic stroke risk, genetically proxied BMI with levels of coagulation factors, and genetically proxied levels of coagulation factors with cardioembolic stroke risk. These estimates were carried forward to mediation and sensitivity analyses.
RESULTS
A 1-SD increase in genetically proxied BMI associated with increased cardioembolic stroke risk (OR of cardioembolic stroke per 1-SD of BMI 1.20, 95% CI 1.08-1.33, = 8.65 × 10) with similar findings in statistical sensitivity analyses more robust to the inclusion of pleiotropic variants. Genetically proxied BMI was further associated with increased levels of Factor VII, Factor Xa, Factor XI, and Protein S (all < 5.9 × 10). Of these factors, genetically proxied levels of Factor XI were associated with cardioembolic stroke risk (OR of cardioembolic stroke per 1-SD increase in Factor XI levels 1.32, 1.19-1.46, = 6.18 × 10). The mediated effect of genetically proxied BMI through Factor XI accounted for 26% (6%-49%) of the total effect of BMI on cardioembolic stroke.
DISCUSSION
Human genetic data support increased levels of Factor XI as a mechanistic explanation for how obesity increases cardioembolic stroke risk. The clinical relevance of this association warrants further investigation within ongoing clinical trials of Factor XI inhibition.
Topics: Humans; Mendelian Randomization Analysis; Obesity; Body Mass Index; Thrombophilia; Stroke; Female; Blood Coagulation Factors; Male; Risk Factors; Embolic Stroke
PubMed: 38861688
DOI: 10.1212/WNL.0000000000209431 -
The Journal of Pharmacology and... Jun 2024Interleukin (IL)-33 has been shown to centrally regulate, among other processes, inflammation and fibrosis. Both intracellular full-length (FLIL33) precursor and...
Interleukin (IL)-33 has been shown to centrally regulate, among other processes, inflammation and fibrosis. Both intracellular full-length (FLIL33) precursor and extracellular mature cytokine (MIL33) forms exert such regulation, albeit differentially. Drug development efforts to target the IL-33 pathway have focused mostly on MIL33 and its specific cell-surface receptor, ST2, with limited attempts to negotiate the pathophysiological contributions from FLIL33. Furthermore, even a successful strategy for targeting MIL33 effects would arguably benefit from a simultaneous attenuation of the levels of FLIL33, which remains the continuous source of MIL33 supply. We therefore sought to develop an approach to depleting FLIL33 protein levels. We previously reported that the steady-state levels of FLIL33 are controlled in part through its proteasomal degradation and that such regulation can be mapped to a segment in the N-terminal portion of FLIL33. We hypothesized that disruption of this regulation would lead to a decrease in FLIL33 levels, thus inducing a beneficial therapeutic effect in an IL-33-dependent pathology. To test this hypothesis, we designed and tested cell-permeable decoy peptides (CPDPs) which mimic the target N-terminal FLIL33 region. We argued that such mimic peptides would compete with FLIL33 for the components of the native FLIL33 production and maintenance molecular machinery. Administered in the therapeutic regimen to bleomycin-challenged mice, the tested CPDPs alleviated the overall severity of the disease by restoring body weight loss and attenuating accumulation of collagen in the lungs. This proof-of-principle study lays the foundation for future work towards the development of this prospective therapeutic approach. An antifibrotic therapeutic approach is proposed and preclinically tested in mice in vivo based on targeting the full-length IL-33 precursor protein. Peptide fusion constructs consisted of a cell-permeable sequence fused with a sequence mimicking an N-terminal segment of IL-33 precursor that is responsible for this protein's stability. Systemic administration of such peptides to mice in either the acute intratracheal or chronic systemic bleomycin challenge models leads to a decrease in the bleomycin-induced elevations of pulmonary IL-33 and collagen.
PubMed: 38858092
DOI: 10.1124/jpet.123.002050 -
Methods in Molecular Biology (Clifton,... 2024Patch-clamp technique provides a unique possibility to record the ion channels' activity. This method enables tracking the changes in their functional states at...
Patch-clamp technique provides a unique possibility to record the ion channels' activity. This method enables tracking the changes in their functional states at controlled conditions on a real-time scale. Kinetic parameters evaluated for the patch-clamp signals form the fundamentals of electrophysiological characteristics of the channel functioning. Nevertheless, the noisy series of ionic currents flowing through the channel protein(s) seem to be bountiful of information, and the standard data processing techniques likely unravel only its part. Rapid development of artificial intelligence (AI) techniques, especially machine learning (ML), gives new prospects for whole channelology. Here we consider the question of the AI applications in the patch-clamp signal analysis. It turns out that the AI methods may not only enable for automatizing of signal analysis, but also they can be used in finding inherent patterns of channel gating and allow the researchers to uncover the details of gating machinery, which had been never considered before. In this work, we outline the currently known AI methods that turned out to be utilizable and useful in the analysis of patch-clamp signals. This chapter can be considered an introductory guide to the application of AI methods in the analysis of the time series of channel currents (together with its advantages, disadvantages, and limitations), but we also propose new possible directions in this field.
Topics: Patch-Clamp Techniques; Machine Learning; Ion Channels; Humans; Ion Channel Gating; Animals
PubMed: 38856906
DOI: 10.1007/978-1-0716-3818-7_15 -
Research and Practice in Thrombosis and... May 2024Here, we present a series of illustrated capsules from the State of the Art (SOA) speakers at the 2024 International Society on Thrombosis and Haemostasis Congress in...
Here, we present a series of illustrated capsules from the State of the Art (SOA) speakers at the 2024 International Society on Thrombosis and Haemostasis Congress in Bangkok, Thailand. This year's Congress marks the first time that the International Society on Thrombosis and Haemostasis has held its flagship scientific meeting in Southeast Asia and is the first to be organized by an international Planning Committee. The Bangkok program will feature innovative science and clinical updates from around the world, reflecting the diversity and multidisciplinary growth of our field. In these illustrated SOA capsules, you will find an exploration of novel models of thrombosis and bleeding and biomaterial discoveries that can trigger or block coagulation. Thromboinflammation is now understood to drive many disease states, and the SOA speakers cover cellular and coagulation responses to COVID-19 and other infections. The theme of crosstalk between coagulation and inflammation expands with capsules on protein S signaling, complement, and fibrinolytic inhibitors. Novel agents for hemophilia and thrombosis prevention are introduced. Challenging clinical conditions are also covered, such as inherited platelet disorders and antiphospholipid antibody syndrome. The scientific program in Bangkok will also showcase the work of clinicians and scientists from all parts of the world and chronicle real-world challenges. For example, 2 SOA capsules address the diagnosis and management of von Willebrand disease in low-income settings. Take some time to browse through these short illustrated reviews; we're sure that you'll be entertained, educated, and inspired to further explore the world of thrombosis and hemostasis.
PubMed: 38854821
DOI: 10.1016/j.rpth.2024.102432 -
Computational Biology and Chemistry Jun 2024Understanding the mechanisms underlying interactions between drugs and target proteins is critical for drug discovery. In our earlier studies, we introduced the...
Understanding the mechanisms underlying interactions between drugs and target proteins is critical for drug discovery. In our earlier studies, we introduced the Triangular Spatial Relationship (TSR)-based algorithm, which enables the representation of a protein's 3D structure as a vector of integers (TSR keys). These TSR keys correspond to substructures of the 3D structure of a protein and are computed based on the triangles constructed by all possible triples of C atoms within the protein. In this study, we report on a new TSR-based algorithm for probing drug and target interactions. Specifically, we have extended the previous algorithm in three novel directions: TSR keys for representing the 3D structure of a drug or a ligand, cross TSR keys between drugs and their targets and intra-residual TSR keys for phosphorylated amino acids. The outcomes illustrate the key contributions as follows: (i) The TSR-based method, which uses the TSR keys as features, is unique in its capability to interpret hierarchical relationships of drugs as well as drug - target complexes using common and specific TSR keys. (ii) The method can distinguish not only the binding sites from the rest of the protein structures, but also the binding sites of primary targets from those of off-targets. (iii) The method has the potential to correlate the 3D structures of drugs with their functions. (iv) Representation of 3D structures by TSR keys has its unique advantage in terms of ease of making searching for similar substructures across structure datasets easier. In summary, this study presents a novel computational methodology, with significant advantages, for providing insights into the mechanism underlying drug and target interactions.
PubMed: 38852360
DOI: 10.1016/j.compbiolchem.2024.108117 -
Biochemistry and Molecular Biology... Jun 2024Green fluorescent protein has long been a favorite protein for demonstrating protein purification in the biochemistry lab course. The protein's vivid green color helps...
Green, yellow, or cyan? Introduction of color change mutations into a green thermostable fluorescent protein and characterization during an introduction to biochemistry lab course.
Green fluorescent protein has long been a favorite protein for demonstrating protein purification in the biochemistry lab course. The protein's vivid green color helps demonstrate to students the concept(s) behind affinity or ion exchange chromatography. We designed a series of introduction to biochemistry labs utilizing a thermostable green protein (TGP-E) engineered to have unusually high thermostability. This protein allows students to proceed through purification and characterization without the need to keep protein samples on ice. The 5-week lab series begins with an introduction to molecular biology techniques during weeks 1 and 2, where site-directed mutagenesis is used introduce, a single nucleotide change that shifts the fluorescent spectra of TGP-E to either cyan (CTP-E) or yellow (YTP-E). Students identify successful mutagenesis reaction by the color of a small expression sample after induction with IPTG. Next, students purify either the TGP-E (control-typically one group volunteers), YTP-E, or CTP-E protein as a 1-week lab. During the following week's lab, students run SDS-PAGE to verify protein purity, bicinchoninic acid assay to quantify protein yield, and absorbance and fluorescence spectra to characterize their protein's fluorescent character. The final lab in the series investigates the thermostability of YTP-E and CTP-E compared with TGP-E using a fluorescence plate reader. This 5-week series of experiments provide students with experience in several key biochemistry techniques and allows the students to compare properties of mutations. At the end of the course, the students will write a research report and give a short presentation over their results.
PubMed: 38850239
DOI: 10.1002/bmb.21841 -
Journal of Molecular Biology Jul 2024The heat shock response (HSR) is a gene regulatory program controlling expression of molecular chaperones implicated in aging, cancer, and neurodegenerative disease.... (Review)
Review
The heat shock response (HSR) is a gene regulatory program controlling expression of molecular chaperones implicated in aging, cancer, and neurodegenerative disease. Long presumed to be activated by toxic protein aggregates, recent work suggests a new functional paradigm for the HSR in yeast. Rather than toxic aggregates, adaptive biomolecular condensates comprised of orphan ribosomal proteins (oRP) and stress granule components have been shown to be physiological chaperone clients. By titrating away the chaperones Sis1 and Hsp70 from the transcription factor Hsf1, these condensates activate the HSR. Upon release from Hsp70, Hsf1 forms spatially distinct transcriptional condensates that drive high expression of HSR genes. In this manner, the negative feedback loop controlling HSR activity - in which Hsf1 induces Hsp70 expression and Hsp70 represses Hsf1 activity - is embedded in the biophysics of the system. By analogy to phosphorylation cascades that transmit information via the dynamic activity of kinases, we propose that the HSR is organized as a condensate cascade that transmits information via the localized activity of molecular chaperones.
Topics: Heat-Shock Response; Saccharomyces cerevisiae Proteins; Transcription Factors; HSP70 Heat-Shock Proteins; Molecular Chaperones; Saccharomyces cerevisiae; Heat-Shock Proteins; DNA-Binding Proteins; Biomolecular Condensates; Ribosomal Proteins; Heat Shock Transcription Factors; Phosphorylation
PubMed: 38848866
DOI: 10.1016/j.jmb.2024.168642 -
Molecular Cell Jun 2024In response to stress, eukaryotes activate the integrated stress response (ISR) via phosphorylation of eIF2α to promote the translation of pro-survival effector genes,...
In response to stress, eukaryotes activate the integrated stress response (ISR) via phosphorylation of eIF2α to promote the translation of pro-survival effector genes, such as GCN4 in yeast. Complementing the ISR is the target of rapamycin (TOR) pathway, which regulates eIF4E function. Here, we probe translational control in the absence of eIF4E in Saccharomyces cerevisiae. Intriguingly, we find that loss of eIF4E leads to de-repression of GCN4 translation. In addition, we find that de-repression of GCN4 translation is accompanied by neither eIF2α phosphorylation nor reduction in initiator ternary complex (TC). Our data suggest that when eIF4E levels are depleted, GCN4 translation is de-repressed via a unique mechanism that may involve faster scanning by the small ribosome subunit due to increased local concentration of eIF4A. Overall, our findings suggest that relative levels of eIF4F components are key to ribosome dynamics and may play important roles in translational control of gene expression.
Topics: Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Phosphorylation; Stress, Physiological; Basic-Leucine Zipper Transcription Factors; Eukaryotic Initiation Factor-4F; Protein Biosynthesis; Gene Expression Regulation, Fungal; Eukaryotic Initiation Factor-4E; Eukaryotic Initiation Factor-2; Signal Transduction; Ribosomes; Eukaryotic Initiation Factor-4A
PubMed: 38848692
DOI: 10.1016/j.molcel.2024.04.016 -
Molecular Cell Jun 2024Protein synthesis is metabolically costly and must be tightly coordinated with changing cellular needs and nutrient availability. The cap-binding protein eIF4E makes the...
Protein synthesis is metabolically costly and must be tightly coordinated with changing cellular needs and nutrient availability. The cap-binding protein eIF4E makes the earliest contact between mRNAs and the translation machinery, offering a key regulatory nexus. We acutely depleted this essential protein and found surprisingly modest effects on cell growth and recovery of protein synthesis. Paradoxically, impaired protein biosynthesis upregulated genes involved in the catabolism of aromatic amino acids simultaneously with the induction of the amino acid biosynthetic regulon driven by the integrated stress response factor GCN4. We further identified the translational control of Pho85 cyclin 5 (PCL5), a negative regulator of Gcn4, that provides a consistent protein-to-mRNA ratio under varied translation environments. This regulation depended in part on a uniquely long poly(A) tract in the PCL5 5' UTR and poly(A) binding protein. Collectively, these results highlight how eIF4E connects protein synthesis to metabolic gene regulation, uncovering mechanisms controlling translation during environmental challenges.
Topics: Eukaryotic Initiation Factor-4E; Saccharomyces cerevisiae Proteins; Protein Biosynthesis; Amino Acids; Saccharomyces cerevisiae; Gene Expression Regulation, Fungal; RNA, Messenger; 5' Untranslated Regions; Basic-Leucine Zipper Transcription Factors; Cyclins; Poly(A)-Binding Proteins
PubMed: 38848691
DOI: 10.1016/j.molcel.2024.05.008 -
Science Advances Jun 2024The hierarchical chromatin organization begins with formation of nucleosomes, which fold into chromatin domains punctuated by boundaries and ultimately chromosomes. In a...
The hierarchical chromatin organization begins with formation of nucleosomes, which fold into chromatin domains punctuated by boundaries and ultimately chromosomes. In a hierarchal organization, lower levels shape higher levels. However, the dependence of higher-order 3D chromatin organization on the nucleosome-level organization has not been studied in cells. We investigated the relationship between nucleosome-level organization and higher-order chromatin organization by perturbing nucleosomes across the genome by deleting () and () chromatin remodeling factors in budding yeast. We find that changes in nucleosome-level properties are accompanied by changes in 3D chromatin organization. Short-range chromatin contacts up to a few kilo-base pairs decrease, chromatin domains weaken, and boundary strength decreases. Boundary strength scales with accessibility and moderately with width of nucleosome-depleted region. Change in nucleosome positioning seems to alter the stiffness of chromatin, which can affect formation of chromatin contacts. Our results suggest a biomechanical "bottom-up" mechanism by which nucleosome distribution across genome shapes 3D chromatin organization.
Topics: Nucleosomes; Chromatin Assembly and Disassembly; Chromatin; Genome, Fungal; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; DNA-Binding Proteins; Transcription Factors; Adenosine Triphosphatases
PubMed: 38848364
DOI: 10.1126/sciadv.adn2955