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Bone Aug 2023Chronic kidney disease (CKD) is characterized by secondary hyperparathyroidism and an increased risk of hip fractures predominantly related to cortical porosity....
Chronic kidney disease (CKD) is characterized by secondary hyperparathyroidism and an increased risk of hip fractures predominantly related to cortical porosity. Unfortunately, bone mineral density measurements and high-resolution peripheral computed tomography (HR-pQCT) imaging have shortcomings that limit their utility in these patients. Ultrashort echo time magnetic resonance imaging (UTE-MRI) has the potential to overcome these limitations by providing an alternative assessment of cortical porosity. The goal of the current study was to determine if UTE-MRI could detect changes in porosity in an established rat model of CKD. Cy/+ rats (n = 11), an established animal model of CKD-MBD, and their normal littermates (n = 12) were imaged using microcomputed tomography (microCT) and UTE-MRI at 30 and 35 weeks of age (which approximates late-stage kidney disease in humans). Images were obtained at the distal tibia and the proximal femur. Cortical porosity was assessed using the percent porosity (Pore%) calculated from microCT imaging and the porosity index (PI) calculated from UTE-MRI. Correlations between Pore% and PI were also calculated. Cy/+ rats had higher Pore% than normal rats at both skeletal sites at 35 weeks (tibia = 7.13 % +/- 5.59 % vs. 0.51 % +/- 0.09 %, femur = 19.99 % +/- 7.72 % vs. 2.72 % +/- 0.32 %). They also had greater PI at the distal tibia at 30 weeks of age (0.47 +/- 0.06 vs. 0.40 +/- 0.08). However, Pore% and PI were only correlated in the proximal femur at 35 weeks of age (ρ = 0.929, Spearman). These microCT results are consistent with prior studies in this animal model utilizing microCT imaging. The UTE-MRI results were inconsistent, resulting in variable correlations with microCT imaging, which may be related to suboptimal bound and pore water discrimination at higher magnetic field strengths. Nevertheless, UTE-MRI may still provide an additional clinical tool to assess fracture risk without using ionizing radiation in CKD patients.
Topics: Humans; Animals; Rats; X-Ray Microtomography; Porosity; Cortical Bone; Magnetic Resonance Imaging; Bone Density; Models, Animal; Renal Insufficiency, Chronic; Hip Fractures
PubMed: 37207990
DOI: 10.1016/j.bone.2023.116808 -
ELife Sep 2023During apoptosis, caspases degrade 8 out of ~30 nucleoporins to irreversibly demolish the nuclear pore complex. However, for poorly understood reasons, caspases are also...
During apoptosis, caspases degrade 8 out of ~30 nucleoporins to irreversibly demolish the nuclear pore complex. However, for poorly understood reasons, caspases are also activated during cell differentiation. Here, we show that sublethal activation of caspases during myogenesis results in the transient proteolysis of four peripheral Nups and one transmembrane Nup. 'Trimmed' NPCs become nuclear export-defective, and we identified in an unbiased manner several classes of cytoplasmic, plasma membrane, and mitochondrial proteins that rapidly accumulate in the nucleus. NPC trimming by non-apoptotic caspases was also observed in neurogenesis and endoplasmic reticulum stress. Our results suggest that caspases can reversibly modulate nuclear transport activity, which allows them to function as agents of cell differentiation and adaptation at sublethal levels.
Topics: Caspases; Nuclear Pore; Cell Differentiation; Apoptosis; Endoplasmic Reticulum Stress
PubMed: 37665327
DOI: 10.7554/eLife.89066 -
JACS Au Sep 2023The use of nanopores for the single-molecule sensing of folded proteins and biomacromolecules has recently gained attention. Here, we introduce a simplified synthetic...
The use of nanopores for the single-molecule sensing of folded proteins and biomacromolecules has recently gained attention. Here, we introduce a simplified synthetic α-helical transmembrane pore, pPorA, as a nanoreactor and sensor that exhibits functional versatility comparable to that of engineered protein and DNA nanopores. The pore, built from the assembly of synthetic 40-amino-acid-long peptides, is designed to contain cysteine residues within the lumen and at the pore terminus for site-specific chemical modification probed using single-channel electrical recordings. The reaction of the pore with differently charged activated thiol reagents was studied, wherein positively charged reagents electrophoretically driven into the pore resulted in pore blocking in discrete steps upon covalent bond formation. The asymmetric blockage patterns resulting from cis and trans-side addition of reagents reveal the pore orientation in the lipid membrane. Furthermore, activated PEG thiols covalently blocked the pores over a longer duration in a charge-independent manner, establishing the large diameter and orientation of the formed pores. While the covalent binding of thiol reagents caused a drop in the pore conductance, cationic cyclic octasaccharides produced time-resolved translocation events, confirming the structural flexibility and tunability of the pores. The ability of the pore to accommodate large analytes and the considerable current amplitude variation following bond formation events are promising for developing platforms to resolve multistep chemical reactions at the single-molecule level for applications in synthetic nanobiotechnology.
PubMed: 37772177
DOI: 10.1021/jacsau.3c00221 -
BioRxiv : the Preprint Server For... Aug 2023Hyperpolarization and cyclic-nucleotide (HCN) activated ion channels play a critical role in generating self-propagating action potentials in pacemaking and rhythmic...
Hyperpolarization and cyclic-nucleotide (HCN) activated ion channels play a critical role in generating self-propagating action potentials in pacemaking and rhythmic electrical circuits in the human body. Unlike most voltage-gated ion channels, the HCN channels activate upon membrane hyperpolarization, but the structural mechanisms underlying this gating behavior remain unclear. Here, we present cryo-electron microscopy structures of human HCN1 in Closed, Intermediate, and Open states. Our structures reveal that the inward motion of two gating charges past the charge transfer center (CTC) and concomitant tilting of the S5 helix drives the opening of the central pore. In the intermediate state structure, a single gating charge is positioned below the CTC and the pore appears closed, whereas in the open state structure, both charges move past CTC and the pore is fully open. Remarkably, the downward motion of the voltage sensor is accompanied by progressive unwinding of the inner end of S4 and S5 helices disrupting the tight gating interface that stabilizes the Closed state structure. This "melting" transition at the intracellular gating interface leads to a concerted iris-like displacement of S5 and S6 helices, resulting in pore opening. These findings reveal key structural features that are likely to underlie reversed voltage-dependence of HCN channels.
PubMed: 37645882
DOI: 10.1101/2023.08.17.553623 -
Nucleus (Austin, Tex.) Dec 2024Cellular behavior is continuously influenced by mechanical forces. These forces span the cytoskeleton and reach the nucleus, where they trigger mechanotransduction... (Review)
Review
Cellular behavior is continuously influenced by mechanical forces. These forces span the cytoskeleton and reach the nucleus, where they trigger mechanotransduction pathways that regulate downstream biochemical events. Therefore, the nucleus has emerged as a regulator of cellular response to mechanical stimuli. Cell cycle progression is regulated by cyclin-CDK complexes. Recent studies demonstrated these biochemical pathways are influenced by mechanical signals, highlighting the interdependence of cellular mechanics and cell cycle regulation. In particular, the transition from G2 to mitosis (G2-M) shows significant changes in nuclear structure and organization, ranging from nuclear pore complex (NPC) and nuclear lamina disassembly to chromosome condensation. The remodeling of these mechanically active nuclear components indicates that mitotic entry is particularly sensitive to forces. Here, we address how mechanical forces crosstalk with the nucleus to determine the timing and efficiency of the G2-M transition. Finally, we discuss how the deregulation of nuclear mechanics has consequences for mitosis.
Topics: Mechanotransduction, Cellular; Cell Nucleus; Mitosis; Cytoskeleton; Biophysics
PubMed: 38533923
DOI: 10.1080/19491034.2024.2330947 -
Nature Communications Nov 2023Coronavirus replication is associated with the remodeling of cellular membranes, resulting in the formation of double-membrane vesicles (DMVs). A DMV-spanning pore was...
Coronavirus replication is associated with the remodeling of cellular membranes, resulting in the formation of double-membrane vesicles (DMVs). A DMV-spanning pore was identified as a putative portal for viral RNA. However, the exact components and the structure of the SARS-CoV-2 DMV pore remain to be determined. Here, we investigate the structure of the DMV pore by in situ cryo-electron tomography combined with subtomogram averaging. We identify non-structural protein (nsp) 3 and 4 as minimal components required for the formation of a DMV-spanning pore, which is dependent on nsp3-4 proteolytic cleavage. In addition, we show that Mac2-Mac3-DPUP-Ubl2 domains are critical for nsp3 oligomerization and crown integrity which influences membrane curvature required for biogenesis of DMVs. Altogether, SARS-CoV-2 nsp3-4 have a dual role by driving the biogenesis of replication organelles and assembly of DMV-spanning pores which we propose here to term replicopores.
Topics: Humans; SARS-CoV-2; Virus Replication; Viral Nonstructural Proteins; COVID-19; Organelles
PubMed: 38036567
DOI: 10.1038/s41467-023-43666-5 -
Nanomaterials (Basel, Switzerland) Sep 2023Hyper-crosslinked porous organic nanomaterials, especially the hyper-crosslinked polymers (HCPs), are a unique class of materials that combine the benefits of high... (Review)
Review
Hyper-crosslinked porous organic nanomaterials, especially the hyper-crosslinked polymers (HCPs), are a unique class of materials that combine the benefits of high surface area, porous structure, and good chemical and thermal stability all rolled into one. A wide range of synthetic methods offer an enormous variety of HCPs with different pore structures and morphologies, which has allowed HCPs to be developed for gas adsorption and separations, chemical adsorption and encapsulation, and heterogeneous catalysis. Here, we present a systematic review of recent approaches to pore size modulation and morphological tailoring of HCPs and their applications to catalysis. We mainly compare the effects of pore size modulation and morphological tailoring on catalytic applications, aiming to pave the way for researchers to develop HCPs with an optimal performance for modern applications.
PubMed: 37764543
DOI: 10.3390/nano13182514 -
Current Opinion in Cell Biology Dec 2023At first glance the nucleus is a highly conserved organelle. Overall nuclear morphology, the octagonal nuclear pore complex, the presence of peripheral heterochromatin... (Review)
Review
At first glance the nucleus is a highly conserved organelle. Overall nuclear morphology, the octagonal nuclear pore complex, the presence of peripheral heterochromatin and the nuclear envelope appear near constant features right down to the ultrastructural level. New work is revealing significant compositional divergence within these nuclear structures and their associated functions, likely reflecting adaptations and distinct mechanisms between eukaryotic lineages and especially the trypanosomatids. While many examples of mechanistic divergence currently lack obvious functional interpretations, these studies underscore the malleability of nuclear architecture. I will discuss some recent findings highlighting these facets within trypanosomes, together with the underlying evolutionary framework and make a call for the exploration of nuclear function in non-canonical experimental organisms.
Topics: Nuclear Pore Complex Proteins; Evolution, Molecular; Nuclear Envelope; Nuclear Pore; Trypanosoma; Lamins; Cell Nucleus; Nuclear Lamina
PubMed: 37666024
DOI: 10.1016/j.ceb.2023.102234 -
Nucleus (Austin, Tex.) Dec 2023In adult mammals, many heart muscle cells (cardiomyocytes) are polyploid, do not proliferate (post-mitotic), and, consequently, cannot contribute to heart regeneration.... (Review)
Review
In adult mammals, many heart muscle cells (cardiomyocytes) are polyploid, do not proliferate (post-mitotic), and, consequently, cannot contribute to heart regeneration. In contrast, fetal and neonatal heart muscle cells are diploid, proliferate, and contribute to heart regeneration. We have identified interdependent changes of the nuclear lamina, nuclear pore complexes, and DNA-content (ploidy) in heart muscle cell maturation. These results offer new perspectives on how cells alter their nuclear transport and, with that, their gene regulation in response to extracellular signals. We present how changes of the nuclear lamina alter nuclear pore complexes in heart muscle cells. The consequences of these changes for cellular regeneration and stress response in the heart are discussed.
Topics: Animals; Nuclear Pore; Nuclear Lamina; Ploidies; Cell Differentiation; Lamins; Mammals
PubMed: 37606283
DOI: 10.1080/19491034.2023.2246310 -
Cells May 2024Norbormide (NRB) is a -selective toxicant, which was serendipitously discovered in 1964 and formerly marketed as an eco-friendly rodenticide that was deemed harmless to... (Review)
Review
Norbormide (NRB) is a -selective toxicant, which was serendipitously discovered in 1964 and formerly marketed as an eco-friendly rodenticide that was deemed harmless to non- species. However, due to inconsistent efficacy and the emergence of second-generation anticoagulants, its usage declined, with registration lapsing in 2003. NRBs' lethal action in rats entails irreversible vasoconstriction of peripheral arteries, likely inducing cardiac damage: however, the precise chain of events leading to fatality and the target organs involved remain elusive. This unique contractile effect is exclusive to rat arteries and is induced solely by the endo isomers of NRB, hinting at a specific receptor involvement. Understanding NRB's mechanism of action is crucial for developing species-selective toxicants as alternatives to the broad-spectrum ones currently in use. Recent research efforts have focused on elucidating its cellular mechanisms and sites of action using novel NRB derivatives. The key findings are as follows: NRB selectively opens the rat mitochondrial permeability transition pore, which may be a factor that contributes to its lethal effect; it inhibits rat vascular K channels, which potentially controls its -selective vasoconstricting activity; and it possesses intracellular binding sites in both sensitive and insensitive cells, as revealed by fluorescent derivatives. These studies have led to the development of a prodrug with enhanced pharmacokinetic and toxicological profiles, which is currently undergoing registration as a novel efficacious eco-sustainable -selective toxicant. The NRB-fluorescent derivatives also show promise as non-toxic probes for intracellular organelle labelling. This review documents in more detail these developments and their implications.
Topics: Animals; Rats; Rodenticides; Humans; Vasoconstriction; Mitochondrial Permeability Transition Pore
PubMed: 38727324
DOI: 10.3390/cells13090788