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ELife Apr 2021Diaphanous (DIAPH) three (DIAPH3) is a member of the formin proteins that have the capacity to nucleate and elongate actin filaments and, therefore, to remodel the...
Diaphanous (DIAPH) three (DIAPH3) is a member of the formin proteins that have the capacity to nucleate and elongate actin filaments and, therefore, to remodel the cytoskeleton. DIAPH3 is essential for cytokinesis as its dysfunction impairs the contractile ring and produces multinucleated cells. Here, we report that DIAPH3 localizes at the centrosome during mitosis and regulates the assembly and bipolarity of the mitotic spindle. DIAPH3-deficient cells display disorganized cytoskeleton and multipolar spindles. DIAPH3 deficiency disrupts the expression and/or stability of several proteins including the kinetochore-associated protein SPAG5. DIAPH3 and SPAG5 have similar expression patterns in the developing brain and overlapping subcellular localization during mitosis. Knockdown of SPAG5 phenocopies DIAPH3 deficiency, whereas its overexpression rescues the DIAHP3 knockdown phenotype. Conditional inactivation of in mouse cerebral cortex profoundly disrupts neurogenesis, depleting cortical progenitors and neurons, leading to cortical malformation and autistic-like behavior. Our data uncover the uncharacterized functions of DIAPH3 and provide evidence that this protein belongs to a molecular toolbox that links microtubule dynamics during mitosis to aneuploidy, cell death, fate determination defects, and cortical malformation.
Topics: Animals; Behavior, Animal; Cell Cycle Proteins; Cell Line, Tumor; Cerebral Cortex; Feeding Behavior; Formins; Gene Expression Regulation, Developmental; Genotype; Humans; Locomotion; Maze Learning; Mice; Mice, Knockout; Microtubules; Mitosis; NIH 3T3 Cells; Neurogenesis; Neurons; Phenotype; Social Behavior; Spindle Apparatus
PubMed: 33899739
DOI: 10.7554/eLife.61974 -
Frontiers in Cellular Neuroscience 2020Cellular protein homeostasis, or proteostasis, is indispensable to the survival and function of all cells. Distinct from other cell types, neurons are long-lived,... (Review)
Review
Cellular protein homeostasis, or proteostasis, is indispensable to the survival and function of all cells. Distinct from other cell types, neurons are long-lived, exhibiting architecturally complex and diverse multipolar projection morphologies that can span great distances. These properties present unique demands on proteostatic machinery to dynamically regulate the neuronal proteome in both space and time. Proteostasis is regulated by a distributed network of cellular processes, the proteostasis network (PN), which ensures precise control of protein synthesis, native conformational folding and maintenance, and protein turnover and degradation, collectively safeguarding proteome integrity both under homeostatic conditions and in the contexts of cellular stress, aging, and disease. Dendrites are equipped with distributed cellular machinery for protein synthesis and turnover, including dendritically trafficked ribosomes, chaperones, and autophagosomes. The PN can be subdivided into an adaptive network of three major functional pathways that synergistically govern protein quality control through the action of (1) protein synthesis machinery; (2) maintenance mechanisms including molecular chaperones involved in protein folding; and (3) degradative pathways (, Ubiquitin-Proteasome System (UPS), endolysosomal pathway, and autophagy. Perturbations in any of the three arms of proteostasis can have dramatic effects on neurons, especially on their dendrites, which require tightly controlled homeostasis for proper development and maintenance. Moreover, the critical importance of the PN as a cell surveillance system against protein dyshomeostasis has been highlighted by extensive work demonstrating that the aggregation and/or failure to clear aggregated proteins figures centrally in many neurological disorders. While these studies demonstrate the relevance of derangements in proteostasis to human neurological disease, here we mainly review recent literature on homeostatic developmental roles the PN machinery plays in the establishment, maintenance, and plasticity of stable and dynamic dendritic arbors. Beyond basic housekeeping functions, we consider roles of PN machinery in protein quality control mechanisms linked to dendritic plasticity (, dendritic spine remodeling during LTP); cell-type specificity; dendritic morphogenesis; and dendritic pruning.
PubMed: 33013325
DOI: 10.3389/fncel.2020.00264 -
Cerebral Cortex (New York, N.Y. : 1991) Nov 2023A multi-scale approach elucidated the origin of the error-related-negativity (ERN), with its associated theta-rhythm, and the post-error-positivity (Pe) in macaque...
A multi-scale approach elucidated the origin of the error-related-negativity (ERN), with its associated theta-rhythm, and the post-error-positivity (Pe) in macaque supplementary eye field (SEF). Using biophysical modeling, synaptic inputs to a subpopulation of layer-3 (L3) and layer-5 (L5) pyramidal cells (PCs) were optimized to reproduce error-related spiking modulation and inter-spike intervals. The intrinsic dynamics of dendrites in L5 but not L3 error PCs generate theta rhythmicity with random phases. Saccades synchronized the phases of the theta-rhythm, which was magnified on errors. Contributions from error PCs to the laminar current source density (CSD) observed in SEF were negligible and could not explain the observed association between error-related spiking modulation in L3 PCs and scalp-EEG. CSD from recorded laminar field potentials in SEF was comprised of multipolar components, with monopoles indicating strong electro-diffusion, dendritic/axonal electrotonic current leakage outside SEF, or violations of the model assumptions. Our results also demonstrate the involvement of secondary cortical regions, in addition to SEF, particularly for the later Pe component. The dipolar component from the observed CSD paralleled the ERN dynamics, while the quadrupolar component paralleled the Pe. These results provide the most advanced explanation to date of the cellular mechanisms generating the ERN.
Topics: Animals; Electroencephalography; Theta Rhythm; Pyramidal Cells; Frontal Lobe; Axons; Macaca; Evoked Potentials
PubMed: 37804250
DOI: 10.1093/cercor/bhad367 -
Proceedings of the National Academy of... Mar 2023During mitosis, cells round up and utilize the interphase adhesion sites within the fibrous extracellular matrix (ECM) as guidance cues to orient the mitotic spindles....
During mitosis, cells round up and utilize the interphase adhesion sites within the fibrous extracellular matrix (ECM) as guidance cues to orient the mitotic spindles. Here, using suspended ECM-mimicking nanofiber networks, we explore mitotic outcomes and error distribution for various interphase cell shapes. Elongated cells attached to single fibers through two focal adhesion clusters (FACs) at their extremities result in perfect spherical mitotic cell bodies that undergo significant 3-dimensional (3D) displacement while being held by retraction fibers (RFs). Increasing the number of parallel fibers increases FACs and retraction fiber-driven stability, leading to reduced 3D cell body movement, metaphase plate rotations, increased interkinetochore distances, and significantly faster division times. Interestingly, interphase kite shapes on a crosshatch pattern of four fibers undergo mitosis resembling single-fiber outcomes due to rounded bodies being primarily held in position by RFs from two perpendicular suspended fibers. We develop a cortex-astral microtubule analytical model to capture the retraction fiber dependence of the metaphase plate rotations. We observe that reduced orientational stability, on single fibers, results in increased monopolar mitotic defects, while multipolar defects become dominant as the number of adhered fibers increases. We use a stochastic Monte Carlo simulation of centrosome, chromosome, and membrane interactions to explain the relationship between the observed propensity of monopolar and multipolar defects and the geometry of RFs. Overall, we establish that while bipolar mitosis is robust in fibrous environments, the nature of division errors in fibrous microenvironments is governed by interphase cell shapes and adhesion geometries.
Topics: Cell Nucleus Division; Mitosis; Centrosome; Aircraft; Axons
PubMed: 36848565
DOI: 10.1073/pnas.2120536120 -
Biomedicines Jan 2022Somatostatin (SST) is widely expressed in the brain and plays various, vital roles involved in neuromodulation. The purpose of this study is to characterize the...
Somatostatin (SST) is widely expressed in the brain and plays various, vital roles involved in neuromodulation. The purpose of this study is to characterize the organization of SST neurons in the Mongolian gerbil visual cortex (VC) using immunocytochemistry, quantitative analysis, and confocal microscopy. As a diurnal animal, the Mongolian gerbil provides us with a different perspective to other commonly used nocturnal rodent models. In this study, SST neurons were located in all layers of the VC except in layer I; they were most common in layer V. Most SST neurons were multipolar round/oval or stellate cells. No pyramidal neurons were found. Moreover, 2-color immunofluorescence revealed that only 33.50%, 24.05%, 16.73%, 0%, and 64.57% of SST neurons contained gamma-aminobutyric acid, calbindin-D28K, calretinin, parvalbumin, and calcium/calmodulin-dependent protein kinase II, respectively. In contrast, neuropeptide Y and nitric oxide synthase were abundantly expressed, with 80.07% and 75.41% in SST neurons, respectively. Our immunocytochemical analyses of SST with D and D dopamine receptors and choline acetyltransferase, α and β nicotinic acetylcholine receptors suggest that dopaminergic and cholinergic fibers contact some SST neurons. The results showed some distinguishable features of SST neurons and provided some insight into their afferent circuitry in the gerbil VC. These findings may support future studies investigating the role of SST neurons in visual processing.
PubMed: 35052772
DOI: 10.3390/biomedicines10010092 -
Archives Italiennes de Biologie Dec 2022Spinal cord injury (SCI) is a condition that causes disturbances in normal sensory, motor, and autonomic functions. During SCI, damages occur such as, contusion,...
PURPOSE
Spinal cord injury (SCI) is a condition that causes disturbances in normal sensory, motor, and autonomic functions. During SCI, damages occur such as, contusion, compression, distraction. The aim of this study was to investigate effects of the antioxidative thymoquinone on neuron and glia cells in SCI biochemically, immunohistochemically and ultrastructurally.
METHODS
Male Sprague-Dawley rats were divided into Control, SCI and SCI + Thymoquinone groups. After T10- T11 laminectomy was performed, a metal weight of 15 grams was left down the spinal tube for spinal damage. Immediately after the trauma, the muscles and skin incision were sutured. Thymoquinone was given to the rats by gavage as 30mg/kg/21days. Tissues fixed in 10% formaldehyde, embedded in paraffin wax and immunstained with Caspase-9 and phosphorylated signal transducer and activator of transcription 3 (pSTAT-3) antibodies. Remaining were stored at -80oC for biochemistry. Frozen spinal cord tissues were placed in a phosphate buffer solution and homogenized, centrifuged then used to measure malondialdehyde (MDA) levels, glutathione peroxidase (GSH) and Myeloperoxidase (MPO).
RESULTS
In the SCI group, MDA, MPO, neuronal degeneration, vascular dilatation, inflammation, apoptotic appearance in the nucleus, loss of membrane and cristae in mitochondria, and dilatation in the endoplasmic reticulum were observed due to degeneration in the neuron structure. In the electron microscopic examination of the trauma + thymoquinone group, the membranes of the nuclei of the glial cells were thick and euchromatin, and mitochondria were shortened in length. In the SCI group, pyknosis and apoptotic changes were observed in neuronal structures and nuclei of glia cells in the substantia grisea and substantia alba region, along with positive Caspase-9 activity. An increase in Caspase-9 activity was observed in endothelial cells in blood vessels. In the SCI + thymoquinone group, Caspase-9 expression was positive in some of the cells in the ependymal canal while the cuboidal cells showed a negative Caspase-9 reaction in the majority. A few degenerated neurons in the substantia grisea region showed a positive reaction with Caspase-9. In SCI group, pSTAT-3 expression was positive in degenerated ependymal cells, neuronal structures, and glia cells. pSTAT-3 expression was positive in the endothelium and surrounding aggregated cells of the enlarged blood vessels. In the SCI+ thymoquinone group, pSTAT-3 expression was negative in most of the bipolar and multipolar neuron structures and glial cells in ependymal cells, enlarged blood vessel endothelial cells.
CONCLUSIONS
It has been thought that thymoquinone application in spinal cord injuries may be an antioxidant that can be recommended as an alternative treatment in suppressing the apoptosis of neural cells by significantly reducing the inflammation process.
Topics: Male; Rats; Animals; Caspase 9; Rats, Sprague-Dawley; Endothelial Cells; Spinal Cord Injuries; Antioxidants
PubMed: 36881916
DOI: 10.12871/000398292022344 -
Archives Italiennes de Biologie Dec 2022Spinal cord injury (SCI) causes various neurological consequences that disrupt the structure of axons. The C/EBP Homologous Protein (CHOP) acts in neuronal death by...
PURPOSE
Spinal cord injury (SCI) causes various neurological consequences that disrupt the structure of axons. The C/EBP Homologous Protein (CHOP) acts in neuronal death by apoptosis has been demonstrated in experimental models. Rosmarinic acid (RA) is a phenolic compound used for therapeutic purposes in many diseases. In this study, we investigated the therapeutic effect of Rosmarinic acid application on inflammation and apoptotic development after spinal cord injury.
METHODS
Male Wistar albino rats (n: 24) were assigned to three group: control, SCI and SCI+ RA. All rats were fixed on the operating table after anesthesia, the skin of the thoracic region was opened with a midline incision and the paravertebral muscles were dissected and T10-T11 laminas were exposed. A cylindrical tube of 10 cm length was fixed to the area to be laminectomy. A metal weight of 15 grams was left down the tube. Spinal damage was created, skin incisions were sutured. 50 mg/kg rosmarinic acid was given orally for 7 days after the spinal injury. Spinal tissues were fixed in formaldehyde solution and processed for paraffin wax tissue protocol and 4-5 μm sections were taken with microtome for further immunohistochemical examination. Caspase-12 and CHOP antibodies were applied to sections. Remaining tissues were carried out in glutaraldehyde for the first fixation then in osmium tetroxide for the second. Tissues were kept in pure araldite and thin sections were taken for transmission electron microscope.
RESULTS
Values of malondialdehyde (MDA), myeloperoxidase (MPO), glutathione peroxidase (GSH), neuronal degeneration, vascular dilation, inflammation, CHOP and Caspase-12 expression were increased in SCI group compared to control group. Only glutathione peroxidase content was decreased in SCI group. In SCI group, disruption of basement membrane structure in canalis ependymalis, degeneration in structures of unipolar bipolar and multipolar neurons, and apoptotic changes were seen with increased inflammation in the piamater region and positive CHOP expression in vascular endothelial cells. In SCI+RA group, reorganization of basement membrane pill in canalis ependymalis were observed with mild Caspase-12 activity in some canalis ependymal and glial cells. Also, moderate CHOP expression in multipolar and bipolar neurons and glia cells were observed.
CONCLUSIONS
The application of RA has a significant effect on preventing damage in SCI. It was thought that CHOP and Caspase-12 mediated oxidative stress could be a guide in showing the potential and therapeutic target to stop the apoptotic course after SCI injury.
Topics: Male; Rats; Animals; Rats, Wistar; Caspase 12; Endothelial Cells; Spinal Cord Injuries; Rosmarinic Acid
PubMed: 36881913
DOI: 10.12871/000398292022341 -
Frontiers in Physiology 2022Endomembrane alkali cation (Na, K)/proton (H) exchangers (eNHEs) are increasingly associated with neurological disorders. These eNHEs play integral roles in regulating... (Review)
Review
Endomembrane alkali cation (Na, K)/proton (H) exchangers (eNHEs) are increasingly associated with neurological disorders. These eNHEs play integral roles in regulating the luminal pH, processing, and trafficking of cargo along the secretory (Golgi and post-Golgi vesicles) and endocytic (early, recycling, and late endosomes) pathways, essential regulatory processes vital for neuronal development and plasticity. Given the complex morphology and compartmentalization of multipolar neurons, the contribution of eNHEs in maintaining optimal pH homeostasis and cargo trafficking is especially significant during periods of structural and functional development and remodeling. While the importance of eNHEs has been demonstrated in a variety of non-neuronal cell types, their involvement in neuronal function is less well understood. In this review, we will discuss their emerging roles in excitatory synaptic function, particularly as it pertains to cellular learning and remodeling. We will also explore their connections to neurodevelopmental conditions, including intellectual disability, autism, and attention deficit hyperactivity disorders.
PubMed: 35547574
DOI: 10.3389/fphys.2022.892196