Much is known about the composition and function of the postsynaptic density (PSD), but less is known about its molecular organization. We use EM tomography to delineate the organization of PSDs at glutamatergic synapses in rat hippocampal cultures. The core of the PSD is dominated by vertically oriented filaments, and ImmunoGold labeling shows that PSD-95 is a component of these filaments. Vertical filaments contact two types of transmembrane structures whose sizes and positions match those of glutamate receptors and intermesh with two types of horizontally oriented filaments lying 10–20 nm from the postsynaptic membrane. The longer horizontal filaments link adjacent NMDAR-type structures, whereas the smaller filaments link both NMDA- and AMPAR-type structures. The orthogonal, interlinked scaffold of filaments at the core of the PSD provides a structural basis for understanding dynamic aspects of postsynaptic function.
The output of a rotary motor is characterized by its torque and speed. We measured the torque-speed relationship of the flagellar rotary motor of Escherichia coli by a new method. Small latex spheres were attached to flagellar stubs on cells fixed to the surface of a glass slide. The angular speeds of the spheres were monitored in a weak optical trap by back-focal-plane interferometry in solutions containing different concentrations of the viscous agent Ficoll. Plots of relative torque (viscosity x speed) versus speed were obtained over a wide dynamic range (up to speeds of approximately 300 Hz) at three different temperatures, 22.7, 17.7, and 15.8 degrees C. Results obtained earlier by electrorotation (, Biophys. J. 65:2201-2216) were confirmed. The motor operates in two dynamic regimes. At 23 degrees C, the torque is approximately constant up to a knee speed of nearly 200 Hz, and then it falls rapidly with speed to a zero-torque speed of approximately 350 Hz. In the low-speed regime, torque is insensitive to changes in temperature. In the high-speed regime, it decreases markedly at lower temperature. These results are consistent with models in which torque is generated by a powerstroke mechanism (, Biophys. J. 76:580-587).
PSD-95, a membrane-associated guanylate kinase (MAGUK), is the major scaffolding protein in the excitatory postsynaptic density (PSD) and a potent regulator of synaptic strength. Here we show that PSD-95 is in an extended configuration and positioned into regular arrays of vertical filaments that contact both glutamate receptors and orthogonal horizontal elements layered deep inside the PSD in rat hippocampal spine synapses. RNAi knockdown of PSD-95 leads to loss of entire patches of PSD material, and EM tomography shows that the patchy loss correlates with loss of PSD-95-containing vertical filaments, horizontal elements associated with the vertical filaments, and putative AMPA, but not NMDA receptor type structures. These observations show that the orthogonal molecular scaffold constructed from PSD-95-containing vertical filaments and their associated horizontal elements is essential for sustaining the three dimensional molecular organization of the PSD. Our findings provide a structural basis for understanding the functional role of PSD-95 at the PSD.
The postsynaptic density (PSD)-95 family of membrane-associated guanylate kinases (MAGUKs) are major scaffolding proteins at the PSD in glutamatergic excitatory synapses, where they maintain and modulate synaptic strength. How MAGUKs underlie synaptic strength at the molecular level is still not well understood. Here, we explore the structural and functional roles of MAGUKs at hippocampal excitatory synapses by simultaneous knocking down PSD-95, PSD-93, and synapse-associated protein (SAP)102 and combining electrophysiology and transmission electron microscopic (TEM) tomography imaging to analyze the resulting changes. Acute MAGUK knockdown greatly reduces synaptic transmission mediated by α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPARs) and N-methyl-D-aspartate receptors (NMDARs). This knockdown leads to a significant rise in the number of silent synapses, diminishes the size of PSDs without changes in pre-or postsynaptic membrane, and depletes the number of membraneassociated PSD-95-like vertical filaments and transmembrane structures, identified as AMPARs and NMDARs by EM tomography. The differential distribution of these receptor-like structures and dependence of their abundance on PSD size matches that of AMPARs and NMDARs in the hippocampal synapses. The loss of these structures following MAGUK knockdown tracks the reduction in postsynaptic AMPAR and NMDAR transmission, confirming the structural identities of these two types of receptors. These results demonstrate that MAGUKs are required for anchoring both types of glutamate receptors at the PSD and are consistent with a structural model where MAGUKs, corresponding to membraneassociated vertical filaments, are the essential structural proteins that anchor and organize both types of glutamate receptors and govern the overall molecular organization of the PSD.T he postsynaptic density (PSD) at excitatory glutamatergic synapses, appearing in electron micrographs as a prominent electron-dense thickening lining the postsynaptic membrane (1) is a complex macromolecular machine positioned across from synaptic vesicle release sites at the presynaptic active zone. The PSD clusters and organizes neurotransmitter receptors and signaling molecules at the postsynaptic membrane, transmits and processes synaptic signals, and can undergo structural changes to encode and store information (2-5). Two types of ionotropic glutamate receptors, AMPA receptors (AMPARs) and NMDA receptors (NMDARs), present at PSDs of excitatory synapses (6-10) mediate almost all synaptic transmission in the brain (11). Biochemistry and mass spectrometry of the detergent-extracted cellular fraction of PSDs have additionally identified many proteins associated with AMPAR and NMDAR complexes (12, 13).The membrane-associated guanylate kinases (MAGUKs), a class of abundant scaffold proteins consisting of PSD-95, PSD-93, synapse-associated protein (SAP)102, and SAP97, interact directly with NMDARs (14-18). These MAGUK proteins share conserved modular structures consisting of three...
Postsynaptic densities (PSDs) contain proteins that regulate synaptic transmission. We determined the positions of calcium/calmodulin-dependent protein kinase II (CaMKII) and PSD-95 within the three-dimensional structure of isolated PSDs using immunogold labeling, rotary shadowing, and electron microscopic tomography. The results show that all PSDs contain a central mesh immediately underlying the postsynaptic membrane. Label for PSD-95 is found on both the cytoplasmic and cleft sides of this mesh, averaging 12 nm from the cleft side. All PSDs label for PSD-95. The properties of CaMKII labeling are quite different. Label is virtually absent on the cleft sides of PSDs, but can be heavy on the cytoplasmic side at a mean distance of 25 nm from the cleft. In tomograms, CaMKII holoenzymes can be visualized directly, appearing as labeled, tower-like structures reflecting the 20 nm diameter of the holoenzyme. These towers protrude from the cytoplasmic side of the central mesh. There appears to be a local organization of CaMKII, as judged by fact that the nearest-neighbor distances are nearly invariant over a wide range of labeling density for CaMKII. The average density of CaMKII holoenzymes is highly variable, ranging from zero to values approaching a tightly packed state. This variability is significantly higher than that for PSD-95 and is consistent with an information storage role for CaMKII.
The total molecular mass of individual postsynaptic densities (PSDs) isolated from rat forebrain was measured by scanning transmission EM. PSDs had a mean diameter of 360 nm and molecular mass of 1.10 ؎ 0.36 GDa. Because the mass represents the sum of the molecular masses of all of the molecules comprising a PSD, it becomes possible to derive the number of copies of each protein, once its relative mass contribution is known.
Layer-structured, single phase Bi5Fe0.5Co0.5Ti3O15 ceramics was synthesized following a multicalcination procedure. Magnetic moment increases more than three times by substituting half Fe sites by Co ions. The material exhibits an Aurivillius phase with a four-layer unit cell structure, and presents a remarkable coexistence of ferroelectricity and ferromagnetism above room temperature. The measured 2Pr and 2Mr are 13 μC/cm2 and 7.8 memu/g, respectively. The material’s magnetic behavior below 275 °C is relaxationlike and its magnetic Curie temperature is ∼345 °C.
Limited data are available for clinical characteristics of patients with coronavirus disease 2019 (COVID-19) outside Wuhan. This study aimed to describe the clinical characteristics of COVID-19 and identify the risk factors for severe illness of COVID-19 in Jiangsu province, China. Clinical data of hospitalized COVID-19 patients were retrospectively collected in 8 hospitals from 8 cities of Jiangsu province, China. Clinical findings of COVID-19 patients were described and risk factors for severe illness of COVID-19 were analyzed. By Feb 10, 2020, 202 hospitalized patients with COVID-19 were enrolled. The median age of patients was 44.0 years (interquartile range, 33.0-54.0). 55 (27.2%) patients had comorbidities. At the onset of illness, the common symptoms were fever (156 [77.2%]) and cough (120 [59.4%]). 66 (32.7%) patients had lymphopenia. 193 (95.5%) patients had abnormal radiological findings. 11 (5.4%) patients were admitted to the intensive care unit and none of the patients died. 23 (11.4%) patients had severe illness. Severe illness of COVID-19 was independently associated with body mass index (BMI) � 28 kg/m 2 (odds ratio [OR], 9.219; 95% confidence interval [CI], 2.731 to 31.126; P<0.001) and a known history of type 2 diabetes PLOS NEGLECTED TROPICAL DISEASES PLOS Neglected Tropical Diseases | https://doi.org/10.1371/journal.pntd.
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