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ELife Jun 2024Allosteric cooperativity between ATP and substrates is a prominent characteristic of the cAMP-dependent catalytic subunit of protein kinase A (PKA-C). This long-range...
Allosteric cooperativity between ATP and substrates is a prominent characteristic of the cAMP-dependent catalytic subunit of protein kinase A (PKA-C). This long-range synergistic action is involved in substrate recognition and fidelity, and it may also regulate PKA's association with regulatory subunits and other binding partners. To date, a complete understanding of this intramolecular mechanism is still lacking. Here, we integrated NMR(Nuclear Magnetic Resonance)-restrained molecular dynamics simulations and a Markov State Model to characterize the free energy landscape and conformational transitions of PKA-C. We found that the apoenzyme populates a broad free energy basin featuring a conformational ensemble of the active state of PKA-C (ground state) and other basins with lower populations (excited states). The first excited state corresponds to a previously characterized inactive state of PKA-C with the αC helix swinging outward. The second excited state displays a disrupted hydrophobic packing around the regulatory (R) spine, with a flipped configuration of the F100 and F102 residues at the αC-β4 loop. We validated the second excited state by analyzing the F100A mutant of PKA-C, assessing its structural response to ATP and substrate binding. While PKA-C preserves its catalytic efficiency with Kemptide, this mutation rearranges the αC-β4 loop conformation, interrupting the coupling of the two lobes and abolishing the allosteric binding cooperativity. The highly conserved αC-β4 loop emerges as a pivotal element to control the synergistic binding of nucleotide and substrate, explaining how mutations or insertions near or within this motif affect the function and drug sensitivity in homologous kinases.
Topics: Molecular Dynamics Simulation; Allosteric Regulation; Adenosine Triphosphate; Catalytic Domain; Cyclic AMP-Dependent Protein Kinases; Protein Conformation; Protein Binding; Nucleotides; Substrate Specificity; Cyclic AMP-Dependent Protein Kinase Catalytic Subunits
PubMed: 38913408
DOI: 10.7554/eLife.91506 -
Heliyon Jun 2024Alveolar epithelial barrier integrity is essential for lung homeostasis. Na, K-ATPase β1 subunit (ATP1B1) involves alveolar edema fluid clearance and alveolar...
AIMS
Alveolar epithelial barrier integrity is essential for lung homeostasis. Na, K-ATPase β1 subunit (ATP1B1) involves alveolar edema fluid clearance and alveolar epithelial barrier stability. However, the underlying molecular mechanism of ATP1B1 in alveolar epithelial cells still needs to be understood.
MAIN METHODS
We utilized Co-Immunoprecipitation mass spectrometry proteomic analysis, protein-protein interaction (PPI) analysis, enrichment analysis, and parallel reaction monitoring (PRM) analysis to investigate proteins interacting with ATP1B1 in A549 cells.
KEY FINDINGS
A total of 159 proteins were identified as significant proteins interacting with ATP1B1 in A549 cells. Ribosomal and heat shock proteins were major constituents of the two main functional modules based on the PPI network. Enrichment analysis showed that significant proteins were involved in protein translation, posttranslational processing, and function regulation. Moreover, 10 proteins of interest were verified by PRM, and fold changes in 6 proteins were consistent with proteomics results. Finally, HSP90AB1, EIF4A1, TUBB4B, HSPA8, STAT1, and PLEC were considered candidates for binding to ATP1B1 to function in alveolar epithelial cells.
SIGNIFICANCE
Our study provides new insights into the role of ATP1B1 in alveolar epithelial cells and indicates that six proteins, in particular HSP90AB1, may be key proteins interacting with and regulating ATP1B1, which might be potential targets for the treatment of acute respiratory distress syndrome.
PubMed: 38912441
DOI: 10.1016/j.heliyon.2024.e32579 -
Frontiers in Immunology 2024The Nipah virus (NiV), a highly deadly bat-borne paramyxovirus, poses a substantial threat due to recurrent outbreaks in specific regions, causing severe respiratory and...
The Nipah virus (NiV), a highly deadly bat-borne paramyxovirus, poses a substantial threat due to recurrent outbreaks in specific regions, causing severe respiratory and neurological diseases with high morbidity. Two distinct strains, NiV-Malaysia (NiV-M) and NiV-Bangladesh (NiV-B), contribute to outbreaks in different geographical areas. Currently, there are no commercially licensed vaccines or drugs available for prevention or treatment. In response to this urgent need for protection against NiV and related infections, we developed a novel homotypic virus-like nanoparticle (VLP) vaccine co-displaying NiV attachment glycoproteins (G) from both strains, utilizing the self-assembling properties of ferritin protein. In comparison to the NiV G subunit vaccine, our nanoparticle vaccine elicited significantly higher levels of neutralizing antibodies and provided complete protection against a lethal challenge with NiV infection in Syrian hamsters. Remarkably, the nanoparticle vaccine stimulated the production of antibodies that exhibited superior cross-reactivity to homologous or heterologous . These findings underscore the potential utility of ferritin-based nanoparticle vaccines in providing both broad-spectrum and long-term protection against NiV and emerging zoonotic challenges.
Topics: Animals; Nipah Virus; Henipavirus Infections; Ferritins; Mesocricetus; Antibodies, Viral; Antibodies, Neutralizing; Nanoparticles; Viral Vaccines; Cricetinae; Vaccines, Virus-Like Particle; Female; Humans; Nanovaccines
PubMed: 38911870
DOI: 10.3389/fimmu.2024.1387811 -
Journal of Cancer 2024Actin-related protein 2/3 complex subunit 1A (ARPC1A) is implicated in several cancers due to its critical role in regulating actin polymerization. However, the exact...
Actin-related protein 2/3 complex subunit 1A (ARPC1A) is implicated in several cancers due to its critical role in regulating actin polymerization. However, the exact mechanism of ARPC1A in cancer remains unclear. This study aims to investigate the biological role of ARPC1A in various cancers and the regulatory role of ARPC1A in glioblastoma multiforme (GBM). We analyzed the expression differences, prognostic value, mutations, immune infiltration, immune microenvironment, and single-cell level correlations of ARPC1A in various cancers. Furthermore, we employed gene set enrichment analysis (GSEA) and functional experiments to elucidate the regulatory mechanisms of ARPC1A on GBM. Importantly, we assessed the role of ARPC1A in temozolomide (TMZ) resistance of GBM. ARPC1A expression was up-regulated in most cancer tissues and was associated with poorer prognosis. Genomic mutation analysis revealed that the predominant type of ARPC1A mutation in tumors was amplification. ARPC1A expression was negatively correlated with B-cell and immune scores in most tumors. Both GSEA and single-cell sequencing have revealed that ARPC1A promotes tumor proliferation and epithelial-mesenchymal transition. experiments confirmed that ARPC1A knockdown inhibited the proliferation and metastatic ability of GBM cells. Notably, silencing ARPC1A reduced TMZ resistance in GBM cells. This study highlights the prognostic value of ARPC1A in various tumors and its potential for application in immunotherapy. Meanwhile, the modulation of GBM malignant behavior and TMZ resistance by ARPC1A provides a new approach for personalized and precise treatment of GBM.
PubMed: 38911374
DOI: 10.7150/jca.94552 -
Current Opinion in Microbiology Jun 2024Bacterial microcompartments (BMCs) are polyhedral structures that segregate enzymatic cargo from the cytosol via encapsulation within a protein shell. Unlike other... (Review)
Review
Bacterial microcompartments (BMCs) are polyhedral structures that segregate enzymatic cargo from the cytosol via encapsulation within a protein shell. Unlike other biological polyhedra, such as viral capsids and encapsulins, BMC shells can exhibit a highly advantageous structural and functional plasticity, conforming to a variety of anabolic (CO fixation in carboxysomes) and catabolic (nutrient assimilation in metabolosomes) roles. Consequently, understanding the subunit properties and associated protein-protein interaction processes that guide shell assembly and function is a necessary step to fully harness BMCs as modular, biotechnological nanomachines. Here, we describe the recent insights into the dynamics of structural features of the key BMC domain (Pfam00936)-containing proteins, which serve as a structural template for BMC-H and BMC-T shell building blocks.
PubMed: 38909546
DOI: 10.1016/j.mib.2024.102497 -
Journal For Immunotherapy of Cancer Jun 2024The dynamic interplay between tyrosine kinase inhibitors (TKIs) and the tumor immune microenvironment (TME) plays a crucial role in the therapeutic trajectory of...
BACKGROUND
The dynamic interplay between tyrosine kinase inhibitors (TKIs) and the tumor immune microenvironment (TME) plays a crucial role in the therapeutic trajectory of non-small cell lung cancer (NSCLC). Understanding the functional dynamics and resistance mechanisms of TKIs is essential for advancing the treatment of NSCLC.
METHODS
This study assessed the effects of short-term and long-term TKI treatments on the TME in NSCLC, particularly targeting epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) mutations. We analyzed changes in immune cell composition, cytokine profiles, and key proteins involved in immune evasion, such as laminin subunit γ-2 (LAMC2). We also explored the use of aspirin as an adjunct therapy to modulate the TME and counteract TKI resistance.
RESULTS
Short-term TKI treatment enhanced T cell-mediated tumor clearance, reduced immunosuppressive M2 macrophage infiltration, and downregulated LAMC2 expression. Conversely, long-term TKI treatment fostered an immunosuppressive TME, contributing to drug resistance and promoting immune escape. Differential responses were observed among various oncogenic mutations, with ALK-targeted therapies eliciting a stronger antitumor immune response compared with EGFR-targeted therapies. Notably, we found that aspirin has potential in overcoming TKI resistance by modulating the TME and enhancing T cell-mediated tumor clearance.
CONCLUSIONS
These findings offer new insights into the dynamics of TKI-induced changes in the TME, improving our understanding of NSCLC challenges. The study underscores the critical role of the TME in TKI resistance and suggests that adjunct therapies, like aspirin, may provide new strategies to enhance TKI efficacy and overcome resistance.
Topics: Tumor Microenvironment; Humans; Protein Kinase Inhibitors; Lung Neoplasms; Carcinoma, Non-Small-Cell Lung; Animals; Mice; Drug Resistance, Neoplasm; Female; ErbB Receptors; Cell Line, Tumor; Mutation
PubMed: 38908857
DOI: 10.1136/jitc-2024-009165 -
Immunology Letters Jun 2024Ablation of the immune-specific catalytic subunit Cβ2 of protein kinase A is associated with a proinflammatory phenotype and increased sensitivity to autoimmunity in...
Ablation of the immune-specific catalytic subunit Cβ2 of protein kinase A is associated with a proinflammatory phenotype and increased sensitivity to autoimmunity in mice. Here we show that tumour growth of the adenocarcinoma cell line EO771 in the breast and in the lung after injection into the mammary fat pad and tail vein, respectively, was significantly reduced in mice ablated for Cβ2 compared to wild-type mice. In both cases, the breast and lung tumours showed increased infiltration of immune cells in the mice lacking Cβ2 compared to wild-type mice. Despite this, it appeared that solid tissue- versus intravenously injected EO771 cells evoked differential immune responses. This was reflected by significantly increased levels of splenic proinflammatory immune cells and circulating cytokines in Cβ2 ablated mice carrying breast- but not the lung tumours. Moreover, Cβ2 ablated mice injected with EO771 cells showed increased overall survival compared to wild-type mice. Taken together, our results suggest for a role for immune cell-specific Cβ2 in protecting against tumour growth induced by EO771 cells in mice that is reflected in improved overall survival.
PubMed: 38908524
DOI: 10.1016/j.imlet.2024.106884 -
Medicina 2024Cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS) is a late onset neurodegenerative disorder. Its genetic basis has recently been identified in...
Cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS) is a late onset neurodegenerative disorder. Its genetic basis has recently been identified in the gene encoding a subunit of the Replication Factor C (RFC1). We present the case of a 62-year-old woman who experienced a history of a biphasic presentation of imbalance and gait disorders, with rapid onset of symptoms followed by slow and progressive neurological deterioration. The diagnostic process was challenging, and numerous tests were conducted to rule out acquired and genetic causes of ataxia, leading to a diagnosis of late-onset idiopathic cerebellar ataxia. Subsequently, vestibular function tests identified severe bilateral vestibulopathy. This led to considering CANVAS among the diagnoses, which was ultimately confirmed through genetic testing (biallelic expansion of the pentanucleotide AAGGG in the RFC1 gene). This case highlights the importance of this new described genetic disease and its subacute presentation variant, emphasizing the relevance of objective vestibular function tests in idiopathic ataxias to achieve proper diagnosis and eventual genetic counseling for offspring.
Topics: Humans; Female; Middle Aged; Cerebellar Ataxia; Bilateral Vestibulopathy; Syndrome; Replication Protein C; Vestibular Function Tests
PubMed: 38907973
DOI: No ID Found -
BMC Ophthalmology Jun 2024Sleep deprivation (SD) is a common public health problem that contributes to various physiological disorders and increases the risk of ocular diseases. However, whether...
BACKGROUND
Sleep deprivation (SD) is a common public health problem that contributes to various physiological disorders and increases the risk of ocular diseases. However, whether sleep loss can damage corneal endothelial function remains unclear. This study aimed to determine the effect and possible mechanism of SD on the corneal endothelium.
METHODS
Male C57BL/6J mice were subjected to establish SD models. After 10 days, quantitative RT-PCR (qRT-PCR) and western blot or immunostaining for the expression levels of zonula occludens-1 (ZO-1), ATPase Na+/K + transporting subunit alpha 1 (Atp1a1), and core clock genes in the corneal endothelium were evaluated. Reactive oxygen species staining and mitochondrial abundance characterized the mitochondrial function. The regulatory role of Bmal1 was confirmed by specifically knocking down or overexpressing basic helix-loop-helix ARNT like 1 protein (Bmal1) in vivo. In vitro, a mitochondrial stress test was conducted on cultured human corneal endothelial cells upon Bmal1 knockdown.
RESULTS
SD damaged the barrier and pump functions of mouse corneal endothelium, accompanied by mitochondrial dysfunction. Interestingly, SD dramatically downregulated the core clock gene Bmal1 expression level. Bmal1 knockdown disrupted corneal endothelial function, while overexpression of Bmal1 ameliorated the dysfunction induced by SD. Mitochondrial bioenergetic deficiency mediated by Bmal1 was an underlying mechanism for SD induced corneal endothelial dysfunction.
CONCLUSION
The downregulation of Bmal1 expression caused by SD led to corneal endothelial dysfunction via impairing mitochondrial bioenergetics. Our findings offered insight into how SD impairs the physiological function of the corneal endothelium and expanded the understanding of sleep loss leading to ocular diseases.
Topics: Sleep Deprivation; Animals; Mice, Inbred C57BL; Male; Mice; ARNTL Transcription Factors; Down-Regulation; Endothelium, Corneal; Disease Models, Animal; Cells, Cultured; Mitochondria; Blotting, Western; Gene Expression Regulation
PubMed: 38907352
DOI: 10.1186/s12886-024-03524-4 -
Nature Communications Jun 2024Long-term non-progressors (LTNPs) of HIV-1 infection may provide important insights into mechanisms involved in viral control and pathogenesis. Here, our results suggest...
Long-term non-progressors (LTNPs) of HIV-1 infection may provide important insights into mechanisms involved in viral control and pathogenesis. Here, our results suggest that the ribosomal protein lateral stalk subunit P1 (RPLP1) is expressed at higher levels in LTNPs compared to regular progressors (RPs). Functionally, RPLP1 inhibits transcription of clade B HIV-1 strains by occupying the C/EBPβ binding sites in the viral long terminal repeat (LTR). This interaction requires the α-helixes 2 and 4 domains of RPLP1 and is evaded by HIV-1 group M subtype C and group N, O and P strains that do not require C/EBPβ for transcription. We further demonstrate that HIV-1-induced translocation of RPLP1 from the cytoplasm to the nucleus is essential for antiviral activity. Finally, knock-down of RPLP1 promotes reactivation of latent HIV-1 proviruses. Thus, RPLP1 may play a role in the maintenance of HIV-1 latency and resistance to RPLP1 restriction may contribute to the effective spread of clade C HIV-1 strains.
Topics: HIV-1; Humans; Ribosomal Proteins; HIV Long Terminal Repeat; CCAAT-Enhancer-Binding Protein-beta; HIV Infections; Transcription, Genetic; Protein Binding; Virus Latency; Binding Sites; Gene Expression Regulation, Viral; HEK293 Cells; Cell Nucleus
PubMed: 38906865
DOI: 10.1038/s41467-024-49622-1