Activated Clec4n < sup > hi < /sup > Neutrophils Aggravate Lung Injury in an Endothelial IGFBP7 Dependent Manner
Am J Respir Cell Mol Biol. 2024 Apr 4. doi: 10.1165/rcmb.2024-0017OC. Online ahead of print.ABSTRACTThe role of endothelial cells in Acute lung injury (ALI) has been widely elaborated, but little is known about the role of different subtypes of endothelial cells in ALI. ALI models were established by lipopolysaccharide. Single-cell RNA sequencing was used to identify differential molecules in endothelial subtypes and heterogeneity of lung immune cells. Specific antibodies were used to block IGFBP7 protein, and AAVshIGP7 was used to specifically knock down IGFBP7. Here, we found that Insulin-like growth factor binding prote...
Source: Mol Biol Cell - April 4, 2024 Category: Molecular Biology Authors: Linchao Yu Rui He Daoxin Wang Di Qi Source Type: research
Foxq1 Promotes Alveolar Epithelial Cell Death through Tle1-mediated Inhibition of the NFkappaB Signaling Pathway
Am J Respir Cell Mol Biol. 2024 Apr 4. doi: 10.1165/rcmb.2023-0317OC. Online ahead of print.ABSTRACTAcute lung injury is a common respiratory disease characterized by diffuse alveolar injury and interstitial edema, as well as a hyperinflammatory response, lung cell damage and oxidative stress. Foxq1, a member of the FOX family of transcription factors, is expressed in various tissues, such as the lungs, liver, and kidneys, and contributes to various biological processes, such as stress, metabolism, cell cycle arrest, and aging-related apoptosis. However, the role of Foxq1 in acute lung injury is unknown. We constructed ex ...
Source: Mol Biol Cell - April 4, 2024 Category: Molecular Biology Authors: Xi Zhu Ershi Hua Qifeng Tu Mei Liu Liqin Xu Jian Feng Source Type: research
IL-10 Counteracts IFN- γ to Alleviate Acute Lung Injury in a Viral-Bacterial Superinfection Model
Am J Respir Cell Mol Biol. 2024 Apr 4. doi: 10.1165/rcmb.2023-0437OC. Online ahead of print.ABSTRACTImmune activation is essential for lung control of viral and bacterial infection, but an overwhelming inflammatory response often leads to the onset of acute respiratory distress syndrome (ARDS). Interleukin-10 (IL-10) plays a crucial role in regulating the balance between antimicrobial immunity and immunopathology. In the current study, we have investigated the role of IL-10 in acute lung injury (ALI) induced by influenza A virus (IAV) and methicillin-resistant Staphylococcus aureus (MRSA) coinfection. This unique coinfecti...
Source: Mol Biol Cell - April 4, 2024 Category: Molecular Biology Authors: Michael McKelvey Md Bashir Uddin Sunil Palani Shengjun Shao Keer Sun Source Type: research
Dissecting a Brake for Airway Smooth Muscle Cell Movement
Am J Respir Cell Mol Biol. 2024 Apr 3. doi: 10.1165/rcmb.2024-0120ED. Online ahead of print.NO ABSTRACTPMID:38568237 | DOI:10.1165/rcmb.2024-0120ED (Source: Mol Biol Cell)
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Dale D Tang Source Type: research
Three factors ParA, TipN, and DnaA-mediated chromosome replication initiation are contributors of centromere segregation in Caulobacter crescentus
Mol Biol Cell. 2024 Apr 3:mbcE23120503. doi: 10.1091/mbc.E23-12-0503. Online ahead of print.ABSTRACTThe ability of bacteria to maintain chromosomal integrity throughout their life cycle is crucial for survival. In Caulobacter crescentus, the polar factor TipN has been proposed to be involved with the partitioning system ParABS. Cells with tipN knocked out display subtle segregation defects of the centromere-like region parS. We hypothesized that TipN's role with parS segregation is obscured by other forces that are ParABS-independent. To test our hypothesis, we removed one of those forces - chromosome replication - and ana...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Morgan Letzkus Corey Trela Paola E Mera Source Type: research
The MBO2/FAP58 heterodimer stabilizes assembly of inner arm dynein < em > b < /em > and reveals axoneme asymmetries involved in ciliary waveform
Mol Biol Cell. 2024 Apr 3:mbcE23110439. doi: 10.1091/mbc.E23-11-0439. Online ahead of print.ABSTRACTCilia generate three-dimensional waveforms required for cell motility and transport of fluid, mucus, and particles over the cell surface. This movement is driven by multiple dynein motors attached to nine outer doublet microtubules that form the axoneme. The outer and inner arm dyneins are organized into 96 nm repeats tandemly arrayed along the length of the doublets. Motility is regulated in part by projections from the two central pair microtubules that contact radial spokes located near the base of the inner dynein arms i...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Gang Fu Katherine Augspurger Jason Sakizadeh Jaimee Reck Raqual Bower Douglas Tritschler Long Gui Daniela Nicastro Mary E Porter Source Type: research
Buffer Choice and pH Strongly Influence Phase Separation of SARS-CoV-2 Nucleocapsid with RNA
Mol Biol Cell. 2024 Apr 3:mbcE23120500. doi: 10.1091/mbc.E23-12-0500. Online ahead of print.ABSTRACTThe SARS-CoV-2 nucleocapsid (N) protein is crucial for virus replication and genome packaging. N protein forms biomolecular condensates both in vitro and in vivo in a process known as liquid-liquid phase separation (LLPS), but the exact factors regulating LLPS of N protein are not fully understood. Here, we show that pH and buffer choice have a profound impact on LLPS of N protein. The degree of phase separation is highly dependent on the pH of the solution, which is correlated with histidine protonation in N protein. Specif...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Nina C Kathe Mihajlo Novakovic Fr édéric H-T Allain Source Type: research
An antitumor fungal polysaccharide from Fomitopsis officinalis by activating immunity and inhibiting angiogenesis
Int J Biol Macromol. 2024 Apr 1:131320. doi: 10.1016/j.ijbiomac.2024.131320. Online ahead of print.ABSTRACTMacrofungi, a class of unique natural resources, are gaining popularity owing to their potential therapeutic benefits and edibility. From Fomitopsis officinalis, a medicinal macrofungus with anticancer activity, a homogeneous heteropolysaccharide (FOBP50-1) with a molecular weight of 2.21 × 104 g/mol has been extracted and purified. FOBP50-1 was found to be composed of 3-O-methylfucose, fucose, mannose, glucose, and galactose with a ratio of 1: 6.5: 4.4: 8.1: 18.2. The sugar fragments and structure of FOBP50-1 were i...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Yongye Shen Jiantong Hou Wenhui Liu Zhen Lin Lingling Ma Jing Xu Yuanqiang Guo Source Type: research
Crystallographic Studies of KRAS in Complex with Small Molecules and RAS-Binding Proteins
Methods Mol Biol. 2024;2797:47-65. doi: 10.1007/978-1-0716-3822-4_5.ABSTRACTRAS proteins play a vital role in regulating downstream signaling and essential cellular processes, positioning them as key players in normal cellular physiology and disease development. Among the various isoforms of RAS, KRAS stands out as one of the most frequently mutated genes in human cancer. The prevalence of RAS mutations in cancer often involves single amino acid substitutions at codons 12, 13, or 61. These mutations disrupt the RAS protein's inherent ability to transition between its active and inactive states, resulting in a constant acti...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Albert H Chan Dhirendra K Simanshu Source Type: research
Profiling Complex RAS-Effector Interactions Using NMR Spectroscopy
Methods Mol Biol. 2024;2797:195-209. doi: 10.1007/978-1-0716-3822-4_14.ABSTRACTKnowledge of how effectors interact with RAS GTPases is key to understanding how these switch-like proteins function in cells. Effectors bind specifically to GTP-loaded RAS using RAS association (RA) or RAS binding domains (RBDs) that show wide-ranging affinities and thermodynamic characteristics. Both normal development and RAS-induced tumorigenesis depend on multiple distinct effector proteins that are frequently co-expressed and co-localized, suggesting an antagonistic nature to signaling whereby multiple proteins compete for a limited pool o...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Regina Strakhova Matthew J Smith Source Type: research
Probing RAS Function Using Monobody and NanoBiT Technologies
Methods Mol Biol. 2024;2797:211-225. doi: 10.1007/978-1-0716-3822-4_15.ABSTRACTMissense mutations in the RAS family of oncogenes (HRAS, KRAS, and NRAS) are present in approximately 20% of human cancers, making RAS a valuable therapeutic target (Prior et al., Cancer Res 80:2969-2974, 2020). Although decades of research efforts to develop therapeutic inhibitors of RAS were unsuccessful, there has been success in recent years with the entrance of FDA-approved KRASG12C-specific inhibitors to the clinic (Skoulidis et al., N Engl J Med 384:2371-2381, 2021; Jänne et al., N Engl J Med 387:120-131, 2022). Additionally, KRASG12D-sp...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Michael Whaby Rakesh Sathish Nair John P O'Bryan Source Type: research
Real-Time Monitoring of RAS Activity Using In Vitro and In-Cell NMR Spectroscopy
Methods Mol Biol. 2024;2797:237-252. doi: 10.1007/978-1-0716-3822-4_17.ABSTRACTThe activation level of RAS can be determined by GTP hydrolysis rate (khy) and GDP-GTP exchange rates (kex). Either impaired GTP hydrolysis or enhanced GDP-GTP exchange causes the aberrant activation of RAS in oncogenic mutants. Therefore, it is important to quantify the khy and kex for understanding the mechanisms of RAS oncogenesis and drug development. Conventional methods have individually measured the kex and khy of RAS. However, within the intracellular environment, GTP hydrolysis and GDP-GTP exchange reactions occur simultaneously under c...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Qingci Zhao Ichio Shimada Noritaka Nishida Source Type: research
FLIM-FRET Protein-Protein Interaction Assay
In this study, we present a step-by-step experimental protocol for applying FLIM-FRET to investigate protein-protein interactions involving various RAS isoforms and RAS effectors at the live cell's plasma membrane. By utilizing the FRET pair comprising enhanced green fluorescent protein (eGFP) and fluorescent mCherry, we demonstrate that the proximity and possible nanoclustering of eGFP-tagged KRAS4b G12D and mCherry-tagged KRAS4b WT led to a reduction in the donor eGFP's fluorescence lifetime. The donor lifetime of eGFP-tagged KRAS decreases even further when treated with a dimer-inducing small molecule, or in the presenc...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Pedro Andrade Bonilla Rebika Shrestha Source Type: research
High-Throughput Cell-Based Screening of Small Molecule KRAS Signaling Inhibitors Using a Homogeneous Time-Resolved Fluorescence (HTRF) Assay
Methods Mol Biol. 2024;2797:271-285. doi: 10.1007/978-1-0716-3822-4_20.ABSTRACTWith recent advances proving that effective inhibition of KRAS is possible, there have been significant efforts made to develop inhibitors of specific mutant alleles. Here we describe a detailed protocol that employs homogeneous time-resolved fluorescence (HTRF) to identify compounds acting on KRAS signaling in malignant cell lines. This method allows for high-throughput, cell-based screens of large compound libraries for the development of RAS-targeted therapeutics.PMID:38570467 | DOI:10.1007/978-1-0716-3822-4_20 (Source: Mol Biol Cell)
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Brian P Smith Megan Rigby Roger Ma Anna E Maciag Source Type: research
FLAG-KRAS4B as a Model System for KRAS4B Proteoform and PTM Evaluation by Mass Spectrometry
Methods Mol Biol. 2024;2797:299-322. doi: 10.1007/978-1-0716-3822-4_22.ABSTRACTPrior analysis of intact and modified protein forms (proteoforms) of KRAS4B isolated from cell lines and tumor samples by top-down mass spectrometry revealed the presence of novel posttranslational modifications (PTMs) and potential evidence of context-specific KRAS4B modifications. However, low endogenous proteoform signal resulted in ineffective characterization, making it difficult to visualize less abundant PTMs or perform follow-up PTM validation using standard proteomic workflows. The NCI RAS Initiative has developed a model system, whereb...
Source: Mol Biol Cell - April 3, 2024 Category: Molecular Biology Authors: Robert A D'Ippolito Grace M Scheidemantle Brian P Smith Katie Powell Scott Eury Abigail Neish Jennifer Mehalko Lauren Beaumont Nicole Fer Vanessa Wall William Burgan Anna E Maciag Dominic Esposito Caroline J DeHart Source Type: research
